Existing Annotations Review

GO Term	Evidence	Action	Reason
GO:0051321 meiotic cell cycle	IEA GO_REF:0000043	MARK AS OVER ANNOTATED	Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "Meiosis"; snapshot-only, removed in the current GOA release. EME1/MUS81 is a structure-specific endonuclease acting on branched DNA in BOTH mitotic/somatic DNA repair and meiotic recombination - it is not meiosis-restricted. Reason: GOA's removal of this annotation was JUSTIFIED. The keyword "Meiosis" reflects the historical "Essential meiotic endonuclease" name (inherited from S. pombe eme1) but mapping it to "meiotic cell cycle" is over-specific for the rice protein. The dominant, experimentally demonstrated role of OsEME1 in rice is somatic DNA-damage repair: oseme1 mutants are hypersensitive to MMS/Zeocin, accumulate gamma-H2AX foci and show cell-cycle arrest in vegetative tissue, with a striped-leaf/chloroplast phenotype [PMID:40333587]. The rice MUS81-EME1 complex contributes only minimally to meiosis - mus81 mutants have normal chiasma numbers and normal crossover designation and the complex acts only on atypical meiotic intermediates [PMID:36495065]. A keyword-based blanket "meiotic cell cycle" term is therefore an over-annotation; the genuine meiotic contribution is more precisely captured by the IBA term "resolution of meiotic recombination intermediates" (GO:0000712), which is retained in current GOA. Supporting Evidence: PMID:36495065 the total chiasma numbers in mus81 mutants were indistinguishable from wild-type. The numbers of HEI10 foci ... in mus81 were also similar to that of wild-type. PMID:40333587 treatment with 75 μg/mL of MMS or 50 μg/mL of Zeocin strongly inhibited root and shoot growth, especially in the mutants PMID:40333587 OsEME1 regulates chloroplast development and division by maintaining the transcription of chloroplast-related genes and nuclear genome integrity
GO:0051301 cell division	IEA GO_REF:0000043	MARK AS OVER ANNOTATED	Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keywords "Cell division" / "Mitosis"; snapshot-only, removed in current GOA. EME1 acts in cell-cycle-coupled DNA repair, but "cell division" per se is not its molecular role. Reason: GOA's removal of this annotation was JUSTIFIED. The keyword mapping produces an overly generic process term. OsEME1 is a DNA-repair endonuclease whose activity is cell-cycle regulated (MUS81-EME1 is activated at G2/M and during the DNA-damage response in other species [PMID:23584455]), and oseme1 mutants show cell-cycle arrest as a CONSEQUENCE of unrepaired DNA damage [PMID:40333587]. The protein does not have a direct role in the cell-division process itself (e.g. cytokinesis, chromosome segregation machinery). "Cell division" is an indirect downstream/contextual term and is better replaced by specific DNA-repair process terms (DNA repair, double-strand break repair) that are retained in current GOA. Supporting Evidence: PMID:40333587 The DNA contents at the S and G2 stages following Zeocin treatment increased in ... cells compared to KY131 PMID:40333587 mutations in OsEME1 led to cell cycle arrest and a DNA damage response
GO:0006310 DNA recombination	IEA GO_REF:0000043	MARK AS OVER ANNOTATED	Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "DNA recombination"; snapshot-only, removed in current GOA. EME1/MUS81 resolves homologous-recombination intermediates, so the essence of the term is biologically correct, but the keyword mapping produces a non-specific parent term. Reason: GOA's removal of this annotation was reasonable and did NOT lose correct biology. OsEME1 genuinely participates in homologous recombination - it binds and cleaves branched HR intermediates (Y12, pre-X12, X12 substrates) in vitro [PMID:40333587] - so "DNA recombination" is not wrong. However, it is a broad parent term, and the specific HR-repair function is better and still represented in current GOA by the IBA terms "double-strand break repair" (GO:0006302) and "resolution of meiotic recombination intermediates" (GO:0000712) and the IEA term "DNA repair" (GO:0006281). A NEW more precise term, "double-strand break repair via homologous recombination" (GO:0000724), is proposed below. Removal of the generic keyword-derived term is therefore acceptable. Supporting Evidence: PMID:40333587 OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which are typical substrates after HR repair of DNA damage file:ORYSJ/EME1/EME1-deep-research-falcon.md rice OsEME1 likely participates in the canonical SSE pathway for processing recombination/replication structures
GO:0004519 endonuclease activity	IEA GO_REF:0000043	MODIFY	Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keywords "Endonuclease"/"Nuclease"; snapshot-only, removed in current GOA. EME1 IS a structure-specific endonuclease (EC 3.1.22.-), and recombinant OsEME1 directly binds and cleaves branched DNA substrates in vitro via its C-terminal ERCC4 domain. Reason: GOA's removal of this annotation was NOT justified - a correct, core molecular-function annotation was lost. EME1 is genuinely an endonuclease: the UniProt RecName is "Crossover junction endonuclease EME1" with EC 3.1.22.-, and Du et al. (2025) showed directly that recombinant MBP-OsEME1 and its C-terminal fragment bind and cleave the branched Y12/pre-X12/X12 substrates, with four conserved residues required for activity [PMID:40333587]. Critically, the current (2026) GOA release contains NO catalytic molecular-function term for OsEME1 at all - only "DNA binding" (GO:0003677). Removing "endonuclease activity" without replacement leaves the gene with no MF describing its core enzymatic activity. The annotation should be retained, but MODIFIED to the more specific and informative term "crossover junction DNA endonuclease activity" (GO:0008821), which precisely captures the EME1/MUS81 structure-specific branched-DNA cleavage activity (see also the NEW entry below). Proposed replacements: crossover junction DNA endonuclease activity Supporting Evidence: PMID:40333587 The major cleavage products were obtained in OsEME1 and OsEME1-C, and their levels gradually increased over time PMID:40333587 OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which are typical substrates after HR repair of DNA damage file:ORYSJ/EME1/EME1-deep-research-falcon.md Purified OsEME1 directly binds and cleaves branched DNA substrates typical of HR intermediates, including Y12, pre-X12, and X12 ... Supports annotation as a crossover-junction/branched-DNA endonuclease involved in HR-mediated DSB repair.
GO:0016787 hydrolase activity	IEA GO_REF:0000043	MARK AS OVER ANNOTATED	Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "Hydrolase"; snapshot-only, removed in current GOA. "Hydrolase activity" is the high-level parent of the gene's true endonuclease (phosphodiester-bond hydrolase) activity. Reason: GOA's removal of this annotation was JUSTIFIED. "Hydrolase activity" is a very broad grouping term; an endonuclease is a hydrolase, so the term is not wrong, but it is uninformative. The correct, specific catalytic function (endonuclease / crossover junction DNA endonuclease activity) is demonstrated experimentally [PMID:40333587] and should be annotated directly (see the MODIFY on GO:0004519 and the NEW GO:0008821 entry). Retaining a top-level "hydrolase activity" term provides no additional information once the specific endonuclease MF is present. Removal of the vague parent is appropriate. Supporting Evidence: PMID:40333587 we measured the endonuclease activity of OsEME1 by performing a nuclease assay, followed by gel electrophoresis file:ORYSJ/EME1/EME1-deep-research-falcon.md the most direct biochemical description is that OsEME1 binds and cleaves branched DNA substrates representing recombination/repair intermediates ... This is an endonucleolytic cleavage of DNA phosphodiester bonds (EC class 3.1.22.-) on structured substrates.
GO:0046872 metal ion binding	IEA GO_REF:0000043	MARK AS OVER ANNOTATED	Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keywords "Metal-binding"/"Magnesium"/"Calcium"; snapshot-only, removed in current GOA. The MUS81-EME1 nuclease reaction is metal-dependent (Mg2+/Ca2+ cofactors). Reason: GOA's removal of this generic keyword-derived term was acceptable. The UniProt entry lists Mg(2+) and Ca(2+) as cofactors (by similarity), and structure-specific nuclease chemistry is divalent-metal-dependent, so the gene does bind metal ions. However, "metal ion binding" is a broad term inferred from a keyword rather than from gene-specific experimental evidence, and no metal-coordinating residues are annotated on the rice protein. If a metal-binding MF were retained it should be the specific "magnesium ion binding" (GO:0000287); but as a free-standing keyword-derived annotation the broad term adds little, so its removal is reasonable. The metal dependence is implicit in the endonuclease MF. Supporting Evidence: PMID:40333587 Eme1 [methyl methanesulfonate 4 (Mms4) in ...] forms a heterodimeric endonuclease with Mus81 ..., cleaving branched DNA substrates such as HJs, 3′-DNA flaps and replication forks file:ORYSJ/EME1/EME1-deep-research-falcon.md activity is influenced by divalent cations (Mg2+/Ca2+ support activity; Mn2+ can increase activity and product diversity).
GO:0031297 replication fork processing	IBA GO_REF:0000033	ACCEPT	Summary: IBA annotation propagated across the EME1/MUS81 phylogenetic group. The MUS81-EME1 complex processes stalled/collapsed replication-fork intermediates - a conserved mitotic function of the complex. Reason: Well supported by conserved biology and consistent with the UniProt FUNCTION statement ("May be required in mitosis for the processing of stalled or collapsed replication fork intermediates"). In Arabidopsis, MUS81 and RECQ4A process recombination-induced aberrant intermediates during replication, and recq4A mus81 double mutants are lethal [PMID:20971895]. Branched fork structures (3'-flaps, replication forks) are canonical MUS81-EME1 substrates and OsEME1 cleaves replication-fork-type Y/X branched substrates in vitro [PMID:40333587]. The IBA annotation is at an appropriate level of specificity. Supporting Evidence: PMID:20971895 RECQ4A and MUS81 are required for processing recombination-induced aberrant intermediates during replication PMID:24008669 RuvC is also known to cut branched DNA intermediates that originate directly from blocked replication forks, targeting them for origin-independent replication restart
GO:0048476 Holliday junction resolvase complex	IBA GO_REF:0000033	ACCEPT	Summary: IBA annotation: EME1 is a subunit of the MUS81-EME1 structure-specific endonuclease, which acts as a Holliday-junction-resolving complex. Confirmed in rice by direct OsEME1-OsMUS81 interaction. Reason: Strongly supported. EME1/MUS81 heterodimers (heterotetramers) are conserved Holliday- junction resolvases across yeast, mammals and plants. In rice, OsEME1 directly interacts with OsMUS81 - shown by yeast two-hybrid, luciferase complementation and BiFC, with the OsEME1 ERCC4 domain and the OsMUS81 HhH motif mediating the interaction [PMID:40333587]. This is a core cellular-component annotation for EME1. Supporting Evidence: PMID:40333587 OsEME1 interacts with OsMUS81 and ... the ERCC4 domain of OsEME1 is required for this interaction PMID:40333587 Homologues of EME1 and MUS81 interact to form heterotetramers in mammals, yeasts and Arabidopsis file:ORYSJ/EME1/EME1-deep-research-falcon.md OsEME1 physically interacts with OsMUS81; interaction depends on the OsEME1 ERCC4 domain and OsMUS81 HhH motif, and interaction signal localizes to nuclei ... Strong evidence that rice OsEME1 functions as the MUS81 partner in a nuclear structure-specific nuclease complex.
GO:0000712 resolution of meiotic recombination intermediates	IBA GO_REF:0000033	KEEP AS NON CORE	Summary: IBA annotation: the MUS81-EME1 complex resolves meiotic recombination intermediates, contributing to a subset of (interference-insensitive) crossovers. In rice this role is minor and limited to atypical meiotic intermediates. Reason: The annotation is correct but reflects a minor, non-core function for the rice protein. Rice MUS81 is required for resolution of ATYPICAL meiotic recombination intermediates (mus81 zep1 and mus81 fancm double mutants show chromosome fragments and bridges) and the FANCM-dependent interference-insensitive extra crossovers require MUS81 for resolution [PMID:36495065, PMID:37632767]. However, rice mus81 single mutants have wild-type chiasma numbers and normal crossover designation, so meiotic crossover resolution is not the dominant function [PMID:36495065]. Given that the demonstrated dominant role of OsEME1 in rice is somatic DSB/replication repair [PMID:40333587], this meiotic-intermediate-resolution term should be retained but classified as non-core. Supporting Evidence: PMID:36495065 MUS81 ... plays a crucial role in the resolution of atypical meiotic intermediates by working together with other anti-crossover factors PMID:37632767 the meiotic extra COs are not marked with HEI10 and require MUS81 resolvase for resolution
GO:0006302 double-strand break repair	IBA GO_REF:0000033	ACCEPT	Summary: IBA annotation: EME1/MUS81 functions in double-strand break repair. This is the core biological process of OsEME1, supported directly by rice mutant phenotypes. Reason: This is a core function and is well supported by direct experimental evidence in rice. oseme1 loss-of-function mutants are hypersensitive to the DNA-damaging agents MMS and Zeocin, accumulate gamma-H2AX foci (a DSB marker) and show S/G2 cell-cycle arrest, and OsEME1 binds and cleaves branched HR-repair intermediates in vitro [PMID:40333587]. In Arabidopsis, MUS81 is required for efficient synthesis-dependent strand annealing (an HR-mediated DSB repair pathway) [PMID:20971895]. The IBA term is at the right level of specificity; a more precise term (GO:0000724, double-strand break repair via homologous recombination) is proposed below. Supporting Evidence: PMID:40333587 The ... and ... mutants exhibited significantly more γ2HAX foci than KY131 with or without Zeocin treatment ... OsEME1 regulates DSB repair and the cell cycle PMID:20971895 MUS81 and RECQ4A are required for efficient synthesis-dependent strand annealing (SDSA) file:ORYSJ/EME1/EME1-deep-research-falcon.md Loss of OsEME1 caused hypersensitivity to DNA-damaging agents ... consistent with a function in HR-mediated repair and/or recovery from replication-associated lesions ... Supports a role in HR-mediated DSB repair and replication-associated genome stability.
GO:0031573 mitotic intra-S DNA damage checkpoint signaling	IBA GO_REF:0000033	KEEP AS NON CORE	Summary: IBA annotation propagated from fission yeast, where Mus81-Eme1 activity is integrated with the intra-S/DNA-damage checkpoint via ATR(Rad3)/CDK phosphorylation of Eme1. Reason: The annotation captures a genuine, conserved regulatory connection but is not a core function and is supported only indirectly for rice. In S. pombe, DNA-damage-induced activation of Mus81-Eme1 requires Cdc2(CDK1)- and Rad3(ATR)-dependent phosphorylation of Eme1 [PMID:23584455]. In rice, oseme1 mutants show DNA-damage-induced cell-cycle arrest and a DNA-damage response [PMID:40333587], consistent with checkpoint coupling, but there is no direct evidence that OsEME1 itself transduces checkpoint signaling - it is a downstream effector (an endonuclease) whose activity is checkpoint- regulated rather than a checkpoint signaling component. Retain as non-core; OsEME1's relationship to the checkpoint is being regulated BY it, not enacting it. Supporting Evidence: PMID:23584455 DNA damage-induced activation of Mus81-Eme1 ... requires both Cdc2(CDK1)- and Rad3(ATR)-dependent phosphorylation of Eme1 PMID:40333587 mutations in ... led to cell cycle arrest and a DNA damage response
GO:0003677 DNA binding	IEA GO_REF:0000002	ACCEPT	Summary: IEA annotation from InterPro (IPR006166, ERCC4 domain). OsEME1 binds DNA directly - confirmed biochemically. Reason: Correct. The ERCC4 domain has nucleic-acid-binding activity, and recombinant OsEME1 and OsEME1-C show major binding shifts on branched Y12 substrates in EMSA, with a measured substrate Kd of ~16 microM by microscale thermophoresis [PMID:40333587]. "DNA binding" is a generic term; the more informative activity is the structure- specific endonuclease MF (see the NEW GO:0008821 entry), but the DNA-binding IEA is not incorrect and can be accepted. Supporting Evidence: PMID:40333587 Major binding shifts were detected in the OsEME1 and OsEME1-C lines but not in the MBP control
GO:0005634 nucleus	IEA GO_REF:0000120	ACCEPT	Summary: IEA annotation for nuclear localization. Directly confirmed in rice by GFP imaging. Reason: Strongly supported by direct experimental evidence: the OsEME1-GFP fusion protein localizes to the nucleus and overlaps with the nuclear marker H2B-mCherry in a transient expression assay [PMID:40333587]. Nuclear localization is consistent with its role as a nuclear DNA-repair endonuclease and with the UniProt subcellular location. Supporting Evidence: PMID:40333587 the OsEME1-GFP fusion protein localized to the nucleus and overlapped closely with the nuclear protein H2B-mCherry file:ORYSJ/EME1/EME1-deep-research-falcon.md Direct evidence from interaction imaging assays indicates that the OsEME1–OsMUS81 interaction occurs in the nucleus, supporting a primary nuclear role in processing DNA repair/recombination intermediates.
GO:0006281 DNA repair	IEA GO_REF:0000002	ACCEPT	Summary: IEA annotation from InterPro (IPR033310, Mms4/EME1/EME2). DNA repair is a core biological process of OsEME1. Reason: Core function, strongly supported by direct rice evidence. oseme1 mutants are hypersensitive to DNA-damaging agents (MMS, Zeocin), accumulate gamma-H2AX foci and show DNA-damage-induced cell-cycle arrest [PMID:40333587]. OsEME1 cleaves branched HR-repair intermediates in vitro. The term is appropriate, though the more specific "double-strand break repair" (GO:0006302, IBA, retained) and the proposed GO:0000724 better capture the HR-specific mechanism. Supporting Evidence: PMID:40333587 we discovered a biochemical role for OsEME1 in HR repair and a biological role for this protein in chloroplast development
GO:0048476 Holliday junction resolvase complex	IEA GO_REF:0000002	ACCEPT	Summary: IEA annotation from InterPro (IPR033310) for the Holliday junction resolvase complex; duplicates the IBA annotation to the same term. Reason: Correct and consistent with the IBA annotation to the same term and with direct evidence that OsEME1 interacts with OsMUS81 to form a structure-specific endonuclease complex [PMID:40333587]. Duplicate annotations with different evidence codes are acceptable; the IEA provides additional computational support for a well-established complex membership. Supporting Evidence: PMID:40333587 OsEME1 interacts with OsMUS81 and ... the ERCC4 domain of OsEME1 is required for this interaction
GO:0009411 response to UV	IEP PMID:12869764 Collection, mapping, and annotation of over 28,000 cDNA clon...	REMOVE	Summary: IEP annotation citing PMID:12869764, which is the rice full-length cDNA consortium paper ("Collection, mapping, and annotation of over 28,000 cDNA clones from japonica rice"). That paper contains no gene-specific UV-response expression experiment. Reason: This annotation is not supported by its cited reference. PMID:12869764 is a large-scale cDNA collection/sequencing/annotation paper; it reports only clone collection, genome mapping and InterPro-based function assignment, with no expression-response assay for this gene. The IEP (Inferred from Expression Pattern) almost certainly derives from the stress-treated source of a cDNA library (library-of-origin metadata), which is not a valid measurement of UV-induced expression and does not demonstrate biological "involvement in" a UV response. Expression presence is not function. While OsEME1 IS a DNA-damage-repair gene (and could plausibly act in UV-DSB repair), the only experimental DNA-damage data [PMID:40333587] used MMS and Zeocin, not UV. This keyword/library-derived IEP should be removed. Supporting Evidence: PMID:12869764 We collected and completely sequenced 28,469 full-length complementary DNA clones from Oryza sativa L. ssp. japonica cv. Nipponbare.
GO:0009644 response to high light intensity	IEP PMID:12869764 Collection, mapping, and annotation of over 28,000 cDNA clon...	REMOVE	Summary: IEP annotation citing PMID:12869764 (rice full-length cDNA collection paper), which contains no gene-specific high-light-response expression experiment for this gene. Reason: The cited reference does not support this annotation. PMID:12869764 is a cDNA collection/annotation paper with no expression-response assay for OsEME1; the IEP is a library-of-origin / keyword artefact, and expression is not function. Notably, OsEME1 high-light sensitivity IS genuine - but it was demonstrated in a different, later paper (Du et al. 2025, PMID:40333587), where oseme1 mutants show severe leaf striping and cell death after high-light treatment. Even there, high-light sensitivity is a downstream consequence of a nuclear genome-maintenance defect (failure to repair oxidative/replication DNA damage), not a primary "response to high light intensity" function of the protein. This particular IEP annotation, as attributed to PMID:12869764, is unsupported and should be removed. Supporting Evidence: PMID:12869764 Through homology searches of publicly available sequence data, we assigned tentative protein functions to 21,596 clones (75.86%).
GO:0010332 response to gamma radiation	IEP PMID:12869764 Collection, mapping, and annotation of over 28,000 cDNA clon...	REMOVE	Summary: IEP annotation citing PMID:12869764 (rice full-length cDNA collection paper), which contains no gene-specific gamma-radiation-response expression experiment. Reason: Not supported by the cited reference. PMID:12869764 reports cDNA clone collection, mapping and annotation, with no gamma-irradiation expression experiment for OsEME1. The IEP reflects library-of-origin metadata rather than a measured expression response, and expression presence is not evidence of biological involvement. The gene's DNA-repair role is real and captured by the DNA-repair / DSB-repair process terms, but this specific "response to gamma radiation" IEP is an over-annotation attributed to a paper that does not test it, and should be removed. Supporting Evidence: PMID:12869764 Mapping of the cDNA clones to genomic DNA revealed that there are 19,000 to 20,500 transcription units in the rice genome.
GO:0008821 crossover junction DNA endonuclease activity	IDA PMID:40333587 ESSENTIAL MEIOTIC ENDONUCLEASE 1 is required for chloroplast...	NEW	Summary: OsEME1 is a structure-specific endonuclease that binds and cleaves branched Holliday-junction-type DNA substrates. This catalytic molecular function is not represented in current GOA and should be added. Reason: The current GOA release has NO catalytic MF term for OsEME1 (only "DNA binding"), and the retired SPKW "endonuclease activity" was the only catalytic annotation lost. Du et al. (2025) demonstrated directly that recombinant OsEME1 / OsEME1-C bind and cleave the branched Y12, pre-X12 and X12 substrates (typical Holliday-junction / crossover-junction structures generated during HR), and that four conserved ERCC4-domain residues are required for activity [PMID:40333587]. The EME1/MUS81 complex is the canonical crossover-junction (Holliday junction) resolvase. "Crossover junction DNA endonuclease activity" (GO:0008821) is the accurate, specific MF and corresponds to the UniProt RecName "Crossover junction endonuclease EME1" and EC 3.1.22.-. IDA is justified by the in vitro nuclease/EMSA assays. Supporting Evidence: PMID:40333587 OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which are typical substrates after HR repair of DNA damage PMID:40333587 these four conserved amino acids are essential for the endonuclease activity of OsEME1 file:ORYSJ/EME1/EME1-deep-research-falcon.md Conserved residues L439, P487, E490, and K494 are important for activity; L439V, E490A, and K494E markedly reduce binding/cleavage ... Identifies functionally critical conserved residues for annotation of catalytic mechanism/structure-function relationships.
GO:0000724 double-strand break repair via homologous recombination	IMP PMID:40333587 ESSENTIAL MEIOTIC ENDONUCLEASE 1 is required for chloroplast...	NEW	Summary: OsEME1 acts in homologous-recombination-mediated repair of DNA double-strand breaks, resolving the branched HR intermediates (Holliday junctions) that arise during repair. Reason: Current GOA captures "double-strand break repair" (GO:0006302, IBA) and "DNA repair" (GO:0006281, IEA) but not the specific HR mechanism. Du et al. (2025) show that oseme1 mutants are hypersensitive to DNA-damaging agents, accumulate gamma-H2AX foci and show DNA-damage-induced cell-cycle arrest, and that OsEME1 binds/cleaves the branched substrates produced after HR repair; the authors explicitly conclude a "biochemical role for OsEME1 in HR repair" [PMID:40333587]. In Arabidopsis, MUS81 is required for synthesis-dependent strand annealing, an HR sub-pathway [PMID:20971895]. GO:0000724 is the precise process term. IMP is justified by the oseme1 loss-of-function DNA-damage-sensitivity phenotypes. Supporting Evidence: PMID:40333587 This study highlights the role of OsEME1 in regulating chloroplast development by modulating homologous recombination repair in response to damage to double-stranded DNA PMID:40333587 The MUS81–EME complex directly binds to and cleaves typical DNA substrates produced after HR repair, a process conserved in yeasts, mammals, dicots and monocots file:ORYSJ/EME1/EME1-deep-research-falcon.md OsEME1 is best annotated as a nuclear, structure-specific endonuclease that cleaves branched/junction DNA substrates consistent with HR/replication intermediates ... rice OsEME1 likely participates in the canonical SSE pathway for processing recombination/replication structures

Core Functions

OsEME1 is the EME1/MMS4-family subunit of a structure-specific DNA endonuclease that, together with MUS81, binds and cleaves branched DNA intermediates - nicked Holliday junctions, crossover junctions, 3'-flaps, D-loops and replication-fork structures. In rice, recombinant OsEME1 (via its C-terminal ERCC4 domain) directly cleaves Holliday-junction-type substrates.

Molecular Function:

crossover junction DNA endonuclease activity

Directly Involved In:

double-strand break repair via homologous recombination

Cellular Locations:

nucleus

In Complex:

Holliday junction resolvase complex

Supporting Evidence:

PMID:40333587
OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which are typical substrates after HR repair of DNA damage
PMID:40333587
OsEME1 interacts with OsMUS81 and ... the ERCC4 domain of OsEME1 is required for this interaction
file:ORYSJ/EME1/EME1-deep-research-falcon.md
OsEME1 is best annotated as a **nuclear, structure-specific endonuclease** that cleaves **branched/junction DNA substrates** consistent with HR/replication intermediates, and that interacts with **OsMUS81** in the nucleus.

OsEME1 maintains nuclear genome stability by resolving homologous-recombination intermediates and processing stalled/collapsed replication forks during somatic DNA repair. Loss of OsEME1 causes hypersensitivity to DNA-damaging agents, gamma-H2AX accumulation and cell-cycle arrest, and - as a downstream consequence of genome- maintenance failure - defective chloroplast development.

Molecular Function:

crossover junction DNA endonuclease activity

Directly Involved In:

double-strand break repair replication fork processing

Cellular Locations:

nucleus

Supporting Evidence:

PMID:40333587
The ... and ... mutants exhibited significantly more γ2HAX foci than KY131 with or without Zeocin treatment ... OsEME1 regulates DSB repair and the cell cycle
PMID:20971895
RECQ4A and MUS81 are required for processing recombination-induced aberrant intermediates during replication
file:ORYSJ/EME1/EME1-deep-research-falcon.md
OsEME1 is required to prevent accumulation of DNA damage and cell-cycle defects and is necessary for normal chloroplast development and photosynthetic performance—likely via a nuclear genome maintenance mechanism and/or maintenance of expression of chloroplast-development regulators.

References

GO_REF:0000002

Gene Ontology annotation through association of InterPro records with GO terms

InterPro-to-GO mappings (IPR006166 ERCC4 domain; IPR033310 Mms4/EME1/EME2) assign DNA binding, DNA repair and Holliday junction resolvase complex to EME1.

GO_REF:0000043

Gene Ontology annotation based on UniProtKB/Swiss-Prot keyword mapping

SwissProt keyword-derived (SPKW) annotations present in the Sept 2025 goa_uniprot_gcrp snapshot but removed from the current GOA release after GOA retired the keyword2GO pipeline for cellular organisms.
For EME1, the keyword "Endonuclease" mapped to a genuinely correct catalytic MF; its removal without replacement left the gene with no catalytic function term.

GO_REF:0000033

Annotation inferences using phylogenetic trees

EME1/MUS81 functions (Holliday junction resolvase complex, replication fork processing, double-strand break repair, meiotic recombination intermediate resolution) are conserved across the PTHR21077 phylogenetic group.

GO_REF:0000120

Combined Automated Annotation using Multiple IEA Methods

Nuclear localization assigned by combined IEA methods; confirmed experimentally by OsEME1-GFP imaging.

PMID:12869764

Collection, mapping, and annotation of over 28,000 cDNA clones from japonica rice.

A large-scale rice full-length cDNA collection, sequencing, genome-mapping and InterPro-based annotation paper.
Contains no gene-specific expression-response experiment for EME1; cannot support IEP annotations for response to UV, high light or gamma radiation.

PMID:40333587

ESSENTIAL MEIOTIC ENDONUCLEASE 1 is required for chloroplast development and DNA repair in rice.

OsEME1 (LOC_Os04g55500) is a single-copy nuclear gene; the OsEME1-GFP fusion localizes to the nucleus.
Recombinant OsEME1 and its C-terminal ERCC4-domain fragment directly bind and cleave branched DNA substrates (Y12, pre-X12, X12) in vitro; four conserved residues are required for activity.
OsEME1 interacts with OsMUS81 (Y2H, luciferase complementation, BiFC); the OsEME1 ERCC4 domain and the OsMUS81 HhH motif mediate the interaction.
oseme1 loss-of-function mutants are hypersensitive to MMS and Zeocin, accumulate gamma-H2AX foci and show S/G2 cell-cycle arrest.
oseme1 mutants have a striped-albino-leaf / defective-chloroplast phenotype enhanced by high light; the chloroplast defect is downstream of nuclear genome-maintenance failure.
OsEME1 functions in homologous-recombination repair of DNA damage.

PMID:36495065

MUS81 is required for atypical recombination intermediate resolution but not crossover designation in rice.

rice mus81 mutants have wild-type chiasma numbers and normal HEI10 foci; MUS81 does not function in crossover designation in rice.
MUS81 resolves atypical meiotic recombination intermediates - mus81 zep1 and mus81 fancm double mutants show chromosome fragments and bridges.

PMID:37632767

FANCM interacts with the MHF1-MHF2 complex to limit crossover frequency during rice meiosis.

FANCM-dependent interference-insensitive extra crossovers in rice fancm mutants are not marked by HEI10 and require MUS81 resolvase for their resolution.

PMID:23584455

Regulation of Mus81-Eme1 Holliday junction resolvase in response to DNA damage.

In fission yeast, DNA-damage-induced activation of Mus81-Eme1 requires both Cdc2(CDK1)- and Rad3(ATR)-dependent phosphorylation of Eme1.
Mus81-Eme1 activation links the DNA-damage response to checkpoint/cell-cycle progression.

PMID:20971895

RAD5A, RECQ4A, and MUS81 have specific functions in homologous recombination and define different pathways of DNA repair in Arabidopsis thaliana.

Arabidopsis MUS81 and RECQ4A process recombination-induced aberrant intermediates during replication; recq4A mus81 double mutants are lethal.
MUS81 is required for efficient synthesis-dependent strand annealing (SDSA), an HR-mediated DSB repair pathway.

PMID:24008669

Resolving branched DNA intermediates with structure-specific nucleases during replication in eukaryotes.

MUS81-EME1, GEN1/Yen1 and SLX1-SLX4 resolve branched DNA intermediates arising from recombination and from blocked/failing replication forks.

PMID:28049740

Resolvase OsGEN1 Mediates DNA Repair by Homologous Recombination.

Yen1/GEN1 resolvases work together with MUS81-EME1 and SLX1-SLX4 in HR-mediated DNA repair to maintain genome stability.
rice osgen1 mutants show reduced chiasma frequency and persistent DSBs in microspores, indicating overlapping resolvase activities in plants.

file:ORYSJ/EME1/EME1-deep-research-falcon.md

Deep-research report (falcon / Edison Scientific Literature) - functional annotation of rice EME1 (Q0J9J6).

Synthesizes the rice-specific Du et al. 2025 study (Plant Biotechnology Journal, DOI 10.1111/pbi.70101, = PMID:40333587) with Arabidopsis and yeast/human MUS81 literature, concluding OsEME1 is best annotated as a nuclear, structure-specific endonuclease that cleaves branched/junction DNA substrates (Y12, pre-X12, X12) consistent with HR/replication intermediates and interacts with OsMUS81 in the nucleus.
Quantifies the rice biochemistry - purified OsEME1 binds the Y12 junction substrate with Kd = 16.3 +/- 2.2 microM (microscale thermophoresis); the C-terminal ERCC4 fragment carries most nuclease activity; conserved residues L439, P487, E490 and K494 are required (L439V/E490A/K494E abolish binding/cleavage).
Independently corroborates the rice mutant phenotypes - oseme1/k48 is hypersensitive to MMS (75 microg/mL) and Zeocin (50 microg/mL), shows S-phase accumulation and gamma-H2AX-like foci, and has a striped/albino chloroplast phenotype (Fv/Fm 0.53 vs 0.72 WT) enhanced by high light, with 970 (normal-light) and 3548 (high-light) DEGs.
Notes the chloroplast defect is most parsimoniously explained by nuclear genome maintenance and/or regulation of nuclear chloroplast-biogenesis genes (OsEME1 cleaves GLK1/GLK2-derived structured DNA in vitro); no evidence of stable chloroplast localization of the OsEME1 protein.
Flags as a caution that in the rice study OsMUS81 bound but did not cleave X-structure substrates while OsEME1 produced cleavage products - deviating from the canonical cross-species view that MUS81 supplies the catalytic site; to be interpreted cautiously and treated as conserved-pathway inference where direct rice data are absent.

Suggested Experiments

Experiment: Reconstitute the OsEME1-OsMUS81 heterodimer from separately expressed, highly purified subunits and compare nuclease activity on Holliday-junction, 3'-flap and replication-fork substrates with each subunit alone, to determine whether OsEME1 is catalytic or regulatory.

Hypothesis: OsEME1 is the regulatory subunit and MUS81 the catalytic subunit, as in yeast and humans; apparent OsEME1-alone activity reflects the heterodimer being the functional unit.

Type: in vitro reconstituted endonuclease assay

Experiment: Generate oseme1 osgen1 double mutants and quantify meiotic chiasmata, chromosome fragmentation, pollen viability and somatic DNA-damage sensitivity.

Hypothesis: OsEME1/OsMUS81 and OsGEN1 act in partially redundant Holliday-junction resolution pathways, so the double mutant shows synthetic genome-instability defects.

Type: genetic epistasis analysis

Experiment: Test oseme1 mutant sensitivity directly to UV-B/UV-C, gamma irradiation and interstrand-crosslinking agents (e.g. mitomycin C) and measure HR-repair efficiency with a chromosomal recombination reporter.

Hypothesis: OsEME1 is required for repair of replication-blocking and double-strand lesions induced by UV, gamma radiation and crosslinkers, consistent with its branched- DNA substrate preference.

Type: genotoxin-sensitivity and recombination-reporter assay

Deep Research

Falcon

(EME1-deep-research-falcon.md)

this with annotations you find in gene/protein databases, but these can be outdated or inaccurate. Falcon Edison Scientific Literature 28 citations 2 artifacts 2026-05-21T10:27:57.229761

Edison artifact artifact-00 (text/markdown)

## Context ID: pqac-00000022 The requested figures showing OsEME1 endonuclease cleavage assays and OsEME1-OsMUS81 interactions were found in Figures 5, 6, and 7 (image/png)

The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.

You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.

We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.

We are interested in where in or outside the cell the gene product carries out its function.

We are also interested in the signaling or biochemical pathways in which the gene functions. We are less interested in broad pleiotropic effects, except where these elucidate the precise role.

Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.

Research report: Functional annotation of rice EME1 (UniProt Q0J9J6; gene EME1; loci Os04g0648700 / LOC_Os04g55500)

0) Target verification (critical disambiguation)

The symbol EME1 is widely used across eukaryotes, but the rice protein specified by UniProt Q0J9J6 corresponds to Oryza sativa subsp. japonica EME1 (also described as OsEME1; “crossover junction endonuclease EME1”), a member of the EME1/MMS4 family with a conserved ERCC4/XPF-like nuclease domain and expected partnership with MUS81. This identity matches the rice gene LOC_Os04g55500, which has been functionally characterized with MUS81 interaction and structure-specific nuclease activity. (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 11-12)

1) Key concepts and definitions (current understanding)

1.1 Structure-selective endonucleases (SSEs)

Structure-selective endonucleases are nucleases that cleave DNA based on DNA structure (e.g., branched intermediates) rather than a strict primary sequence. In eukaryotic homologous recombination (HR) and replication stress responses, SSEs process branched DNA intermediates such as 3′ flaps, replication-fork-like structures, and Holliday junction (HJ)-like X structures, enabling repair completion and chromosome segregation. In plants, MUS81-associated SSE activities are described as recognizing/processing 3′ flaps, replication fork structures, and nicked HJs. (olivier2016thestructurespecificendonucleases pages 1-5)

1.2 MUS81–EME1 (Mms4) complexes

Across eukaryotes, MUS81 forms an SSE complex with a non-identical partner (EME1 in many species; Mms4 in budding yeast). In Arabidopsis, MUS81 forms two distinct complexes with two EME1 paralogs (AtEME1A/B), and both complexes efficiently cleave 3′ flaps and nicked HJs, while intact HJs are cut less efficiently. (geuting2009twodistinctmus81eme1 pages 1-2, geuting2009twodistinctmus81eme1 pages 5-8)

For rice, recent experimental evidence indicates OsEME1 interacts with OsMUS81 and the interaction localizes to the nucleus, consistent with a role in nuclear DNA repair/recombination intermediate processing. (du2025essentialmeioticendonuclease pages 11-12)

1.3 “Crossover junction endonuclease” and the reaction catalyzed

For the rice target, the most direct biochemical description is that OsEME1 binds and cleaves branched DNA substrates representing recombination/repair intermediates—specifically Y-shaped and X-junction-like substrates (Y12, pre-X12, X12) used as model substrates for HR intermediates. This is an endonucleolytic cleavage of DNA phosphodiester bonds (EC class 3.1.22.-) on structured substrates. (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 12-13)

2) Rice OsEME1: biochemical function, substrates, and mechanism

2.1 Substrate specificity (direct evidence in rice)

Du et al. report that purified rice OsEME1 directly binds and cleaves synthetic DNA substrates Y12, pre-X12 and X12 (branched/junction DNA structures used to model HR intermediates). (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 12-13)

A quantitative binding measurement using microscale thermophoresis reported Kd = 16.321 ± 2.163 µM for OsEME1 binding to a Y12 substrate. (du2025essentialmeioticendonuclease pages 7-11)

In addition, OsEME1 bound and cleaved structured DNA fragments derived from rice GLK1 and GLK2 loci in vitro (tested as Y12-like substrates), suggesting it can act on structured DNA within gene-derived contexts in addition to generic model junctions. (du2025essentialmeioticendonuclease pages 7-11)

2.2 Domain-to-function mapping and essential residues

Biochemical truncation analysis indicates that the C-terminus of OsEME1 is responsible for most observed nuclease activity in vitro. (du2025essentialmeioticendonuclease pages 7-11)

Site-directed mutagenesis identified conserved residues important for binding/cleavage phenotypes (e.g., L439, P487, E490, K494). In the reported assays, L439V, E490A, and K494E greatly reduced binding/cleavage, while P487A reduced binding but increased cleavage-product signal, suggesting altered coupling between binding and catalysis. (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 12-13)

2.3 Partnership with MUS81 (direct evidence in rice)

Multiple protein–protein interaction assays support that OsEME1 interacts with OsMUS81, and domain mapping indicates the ERCC4 domain of OsEME1 and the HhH motif region of OsMUS81 contribute to this interaction. The interaction was visualized in the nucleus by BiFC/LUC-based assays. (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 11-12)

Visual evidence: OsEME1 cleavage assays and OsEME1–OsMUS81 interaction/localization are shown in the retrieved figure panels. (du2025essentialmeioticendonuclease media c8ee333c, du2025essentialmeioticendonuclease media 8341a419)

2.4 Note on catalytic contribution within the rice MUS81–EME1 pair

In the same rice study, OsMUS81 was reported to bind X-structure substrates but not be responsible for cleavage under the specific assay conditions, while OsEME1 produced cleavage products; this differs from the canonical cross-species framing where MUS81 supplies the catalytic XPF nuclease active site and EME1 acts as the obligate partner. This discrepancy should be interpreted cautiously (e.g., assay design, protein constructs, or species-specific division of labor). (du2025essentialmeioticendonuclease pages 11-12)

3) Biological roles in rice: DNA repair, cell cycle, and chloroplast development

3.1 DNA damage resistance and DDR activation

Loss of OsEME1 caused hypersensitivity to DNA-damaging agents, including treatments reported as 75 µg/mL MMS or 50 µg/mL Zeocin in seedling assays, with reduced growth compared to wild type. (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 5-7)

OsEME1 mutants also showed evidence of a DNA damage response (e.g., increased γH2AX-like/cH2AX foci) and cell-cycle perturbation (S-phase accumulation reported using EdU/flow cytometry), consistent with a function in HR-mediated repair and/or recovery from replication-associated lesions. (du2025essentialmeioticendonuclease pages 12-13, du2025essentialmeioticendonuclease pages 5-7)

3.2 Chloroplast development and photosynthesis phenotypes

A forward-genetic rice mutant (k48) defective in OsEME1 exhibited striped/albino leaf phenotypes, chloroplast ultrastructural defects, and reduced photosynthetic performance, including decreased maximum PSII quantum yield Fv/Fm = 0.53 vs 0.72 (wild type) under the reported conditions. (du2025essentialmeioticendonuclease pages 2-4)

Transcriptome profiling in the same work showed broad gene-expression effects consistent with chloroplast and repair pathway involvement, including 970 DEGs under normal light and 3548 DEGs under high light. (du2025essentialmeioticendonuclease pages 5-7)

Mechanistically, the authors link OsEME1 nuclease activity and DNA-repair function to maintenance of expression of chloroplast development regulators (e.g., GLK1/GLK2-associated regulatory network), with OsEME1 capable of cleaving GLK1/GLK2-derived structured DNA fragments in vitro. (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 5-7)

4) Subcellular localization (where the gene product acts)

Direct evidence from interaction imaging assays indicates that the OsEME1–OsMUS81 interaction occurs in the nucleus, supporting a primary nuclear role in processing DNA repair/recombination intermediates. (du2025essentialmeioticendonuclease pages 11-12, du2025essentialmeioticendonuclease media 8341a419)

No direct evidence in the retrieved corpus demonstrated stable chloroplast localization of OsEME1 protein; instead, chloroplast developmental phenotypes are most parsimoniously explained by nuclear genome maintenance and/or regulation of nuclear genes required for chloroplast biogenesis, coupled to a DNA-damage response. (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 5-7)

5) Pathway context: homologous recombination, replication stress, and junction processing

5.1 Plant MUS81–EME1 functional archetype (comparative plant evidence)

In Arabidopsis, recombinant MUS81–EME1 complexes efficiently cleave 3′ flaps and nicked HJs, and cleave intact HJs with reduced efficiency; activity is influenced by divalent cations (Mg2+/Ca2+ support activity; Mn2+ can increase activity and product diversity). (geuting2009twodistinctmus81eme1 pages 1-2, geuting2009twodistinctmus81eme1 pages 5-8)

Plant genetic evidence positions MUS81-associated pathways as important for resolving toxic replication intermediates and maintaining genome stability; for example, MUS81 and SEND1 (GEN1-like) have nonredundant roles in telomere stability and replication intermediate processing in Arabidopsis. (olivier2016thestructurespecificendonucleases pages 1-5)

These plant findings support annotating rice OsEME1 as part of a conserved MUS81–EME1 SSE module operating in HR/replication-associated DNA repair, with substrates centered on branched DNA intermediates. (geuting2009twodistinctmus81eme1 pages 1-2, olivier2016thestructurespecificendonucleases pages 1-5)

5.2 Rice MUS81 context supports a conserved partnership model

Rice OsMUS81 is a single-copy gene with XPF-like features, and an alternatively spliced transcript truncates the C-terminus, likely removing an interaction region expected to be needed for association with an EME1/Mms4 partner (thereby plausibly reducing functionality). (mimida2007twoalternativelyspliced pages 2-3)

6) Recent developments (prioritizing 2023–2024)

6.1 2023: MUS81 complexes and DNA–protein crosslink (DPC) repair (mechanistic expansion)

A 2023 biochemical study showed that yeast Mus81–Mms4 and human MUS81 complexes (including MUS81–EME1) can cleave certain DPC-mimicking substrates (e.g., fluorescein/streptavidin adducts) and substrates bearing proteolytically processed TOP1 remnants, but do not cleave DNA bearing native, intact TOP1 covalent complexes—implying MUS81 incision acts after protein adduct trimming/dislodgement. (marini2023mus81cleavestop1derived pages 1-2, marini2023mus81cleavestop1derived pages 4-6)

In Arabidopsis-focused 2023 work, MUS81 is explicitly discussed as part of plant DPC repair capacity and replication-stress intermediate processing, consistent with genetic hypersensitivity phenotypes (e.g., to camptothecin and cisplatin) when MUS81 is defective. (tomastikova2023smc56complexmediatedsumoylation pages 2-4)

Relevance to rice OsEME1: These studies strengthen the contemporary view that MUS81-partnered SSEs act broadly on stalled-fork and branched intermediates and can contribute to DPC lesion processing after prior proteolysis; for rice, this is best treated as conserved-pathway inference, since rice OsEME1 was directly tested on branched substrates but not directly in DPC incision assays in the retrieved literature. (du2025essentialmeioticendonuclease pages 12-13, marini2023mus81cleavestop1derived pages 4-6)

6.2 2024: limited plant MUS81–EME1-specific updates in retrieved corpus

No 2024 plant primary studies directly characterizing MUS81–EME1 biochemical specificity were retrieved here; 2024 articles located were largely meiosis regulation in non-plant systems and are not used for direct annotation claims about rice OsEME1. (geuting2009twodistinctmus81eme1 pages 2-3)

7) Current applications and real-world implementations

Direct 2023–2024 “real-world” deployment literature specifically manipulating rice EME1 for breeding was not retrieved. Nonetheless, two evidence-grounded application directions follow from current knowledge:

1) Stress resilience and productivity under genotoxic conditions: OsEME1 mutants show strong phenotypes under DNA damage and high-light contexts, suggesting that natural variation or engineered tuning of this pathway could impact growth robustness under stresses that elevate DNA damage (e.g., oxidative stress/light stress). This is an implication grounded in the documented mutant phenotypes, not a demonstrated agronomic implementation. (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 5-7)

2) Genome engineering/breeding frameworks leveraging DNA repair and recombination: Reviews on crop improvement emphasize that DNA repair and recombination pathways are central to breeding and genome modification strategies; MUS81–EME1 is a key SSE node in these networks, implying that recombination/repair factors could be future levers for controlling recombination outcomes or maintaining genome stability during editing/regeneration. (verma2020structuralaspectsof pages 11-11)

8) Expert synthesis and interpretation (authoritative analysis)

Most supported primary function in rice: OsEME1 is best annotated as a nuclear, structure-specific endonuclease that cleaves branched/junction DNA substrates consistent with HR/replication intermediates, and that interacts with OsMUS81 in the nucleus. (du2025essentialmeioticendonuclease pages 11-12, du2025essentialmeioticendonuclease pages 12-13)

Biological role: OsEME1 is required to prevent accumulation of DNA damage and cell-cycle defects and is necessary for normal chloroplast development and photosynthetic performance—likely via a nuclear genome maintenance mechanism and/or maintenance of expression of chloroplast-development regulators. (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 5-7)

Cross-species support: Arabidopsis biochemical work provides strong plant-specific precedent that MUS81–EME1 complexes preferentially process 3′ flaps and nicked HJs, reinforcing that rice OsEME1 likely participates in the canonical SSE pathway for processing recombination/replication structures. (geuting2009twodistinctmus81eme1 pages 1-2, geuting2009twodistinctmus81eme1 pages 5-8)

9) Key statistics and data summary

The table below consolidates the main quantitative and assay-backed evidence supporting functional annotation.

Evidence type	Finding	Quantitative details	Experimental system/assay	Biological implication	Source (with year, DOI URL)
Rice OsEME1 identity/function	OsEME1 corresponds to LOC_Os04g55500 and is characterized as a structure-selective endonuclease in rice; the k48 mutant carries a splice-site G→A change causing intron retention and premature termination.	k48 photosystem efficiency reduced: Fv/Fm 0.53 vs 0.72 in wild type under normal-light conditions reported; striped/albino phenotype strongest under high light.	Map-based cloning, complementation, transcript analysis, physiological phenotyping in Oryza sativa japonica.	Confirms that UniProt Q0J9J6/LOC_Os04g55500 encodes rice OsEME1 required for normal chloroplast function and genome maintenance.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 1-2)
Rice OsEME1 biochemical substrate specificity	Purified OsEME1 directly binds and cleaves branched DNA substrates typical of HR intermediates, including Y12, pre-X12, and X12.	Time-dependent cleavage products observed; full-length protein active, with stronger activity from the C-terminal fragment.	In vitro EMSA and endonuclease cleavage assays using recombinant MBP-OsEME1 and MBP-OsEME1-C on synthetic DNA junction substrates.	Supports annotation as a crossover-junction/branched-DNA endonuclease involved in HR-mediated DSB repair.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 12-13, du2025essentialmeioticendonuclease pages 5-7)
Rice OsEME1 domain/activity mapping	The C terminus of OsEME1 is sufficient for major DNA binding/cleavage activity and contains the ERCC4-associated catalytic region implicated in nuclease function.	C-terminal fragment showed major binding and cleavage in vitro relative to other fragments.	Recombinant truncation analysis with DNA binding and cleavage assays.	Indicates catalytic/structure-recognition determinants reside in the conserved ERCC4-family C-terminal region, consistent with UniProt domain annotation.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 12-13)
Rice OsEME1 DNA binding affinity	OsEME1 binds Y12-type DNA substrate with micromolar affinity.	MST equilibrium dissociation constant Kd = 16.321 ± 2.163 µM for Y12 substrate.	Microscale thermophoresis with recombinant OsEME1 and Y12 substrate.	Provides quantitative support for direct recognition of branched repair intermediates.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11)
Rice OsEME1 catalytic residues	Conserved residues L439, P487, E490, and K494 are important for activity; L439V, E490A, and K494E markedly reduce binding/cleavage, whereas P487A lowers binding but can increase cleavage product accumulation.	Qualitative loss/reduction of binding and cleavage for L439V/E490A/K494E; altered coupling of binding versus cleavage for P487A.	Site-directed mutagenesis followed by DNA binding and cleavage assays.	Identifies functionally critical conserved residues for annotation of catalytic mechanism/structure-function relationships.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 12-13)
Rice OsEME1 target-like DNA fragments	OsEME1 binds and cleaves structured DNA fragments derived from rice GLK1 and GLK2 loci in vitro.	GLK1/GLK2-derived Y12 substrates were cleaved; 17 of 83 chloroplast-development DEGs were GLK1/2 targets in transcriptome analysis.	In vitro binding/cleavage assays using GLK1/GLK2-derived substrates; RNA-seq/GO analysis in mutant seedlings.	Suggests a link between DNA structure processing, genome stability, and transcriptional maintenance of chloroplast developmental regulators.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 5-7)
Rice OsEME1 interaction partner/localization	OsEME1 physically interacts with OsMUS81; interaction depends on the OsEME1 ERCC4 domain and OsMUS81 HhH motif, and interaction signal localizes to nuclei.	Interaction detected by Y2H, LCI, and BiFC; domain mapping implicated OsEME1 F4/ERCC4 and OsMUS81 F5/HhH regions.	Yeast two-hybrid, luciferase complementation imaging, bimolecular fluorescence complementation.	Strong evidence that rice OsEME1 functions as the MUS81 partner in a nuclear structure-specific nuclease complex.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 11-12, du2025essentialmeioticendonuclease pages 1-2)
Rice OsEME1 versus OsMUS81 contribution	In the 2025 rice study, OsMUS81 bound X-structure substrates but was not responsible for cleavage of junction DNA substrates under the tested conditions, whereas OsEME1 produced cleavage products.	Qualitative difference: OsMUS81 binding without evident cleavage of X/pre-X in that assay context.	Recombinant protein binding/cleavage assays with X/pre-X substrates.	Suggests OsEME1 may provide key catalytic contribution or alter complex architecture in rice, though this deviates from the canonical cross-species view and should be interpreted cautiously.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 11-12)
Rice chloroplast phenotype	Loss of OsEME1 disrupts chloroplast development and photosynthesis.	Fv/Fm 0.53 in k48 vs 0.72 in wild type; significantly lower chlorophylls and carotenoids reported; 970 DEGs under normal light and 3548 DEGs under high light.	Seedling phenotyping, pigment/photosynthesis measurements, chloroplast ultrastructure, RNA-seq under normal and high light.	Links genome-maintenance nuclease activity to chloroplast biogenesis and light-stress tolerance in rice.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 2-4, du2025essentialmeioticendonuclease pages 5-7)
Rice DNA damage sensitivity	Oseme1 mutants are hypersensitive to DNA-damaging agents MMS and Zeocin.	Treatments reported: 75 µg/mL MMS or 50 µg/mL Zeocin for 5 days; mutants had reduced shoot/root growth versus wild type.	Chemical sensitivity assays on rice seedlings.	Indicates OsEME1 is required for repair of genotoxic lesions and/or recovery from replication-associated DNA damage.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 11-12, du2025essentialmeioticendonuclease pages 5-7)
Rice cell-cycle and DNA-damage markers	OsEME1 deficiency causes cell-cycle disturbance and elevated DNA damage markers.	Increased S-phase cells / S-G2 accumulation; increased cH2AX/γH2AX-like foci after 200 µg/mL Zeocin challenge reported.	EdU incorporation, flow cytometry/ploidy analysis, immunodetection of DNA damage foci.	Supports a role in HR-mediated DSB repair and replication-associated genome stability.	Du et al., 2025, https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 7-11, du2025essentialmeioticendonuclease pages 12-13, du2025essentialmeioticendonuclease pages 5-7)
Rice MUS81 contextual evidence	Rice OsMUS81 is a single-copy XPF-like nuclease-family protein with HhH motifs; an alternatively spliced form lacks a C-terminal interaction region expected for EME1/Mms4 association and is likely nonfunctional.	Full-length protein 660 aa (~73 kDa); truncated splice form 542 aa (~60 kDa); induced by 100 Gy γ-radiation.	cDNA/genome analysis and expression studies in rice.	Provides independent rice-context support for a conserved OsMUS81–OsEME1 partnership in DNA damage responses.	Mimida et al., 2007, https://doi.org/10.1093/pcp/pcm029 (mimida2007twoalternativelyspliced pages 2-3, mimida2007twoalternativelyspliced pages 4-5)
Cross-species inference: Arabidopsis MUS81-EME1 complexes	Arabidopsis MUS81-EME1 complexes process 3′ flaps and nicked Holliday junctions efficiently and intact HJs less efficiently; two EME1 paralogs (EME1A/B) both form active complexes with MUS81.	AtEME1A and AtEME1B share ~63% identity; AtMUS81-EME1B generally showed higher specific activity; cleavage positions mapped ~3–13 nt 5′ of junction; Mg2+ and Ca2+ support activity, Mn2+ enhances it.	Recombinant coexpression/purification and biochemical nuclease assays in Arabidopsis proteins.	Reinforces the canonical plant annotation of EME1 as the MUS81 partner for cleavage of branched HR/replication intermediates.	Geuting et al., 2009, https://doi.org/10.1104/pp.109.136846 (geuting2009twodistinctmus81eme1 pages 1-2, geuting2009twodistinctmus81eme1 pages 2-3, geuting2009twodistinctmus81eme1 pages 5-8)
Cross-species inference: plant DNA repair/telomere stability	Arabidopsis genetics place MUS81 in repair of stalled/collapsed replication intermediates, crosslink repair, and telomere stability; MUS81 and SEND1 together are essential for resolving toxic replication intermediates.	mus81 mutants are hypersensitive to DNA damage; double loss with SEND1 causes severe developmental and genome-instability phenotypes.	Arabidopsis mutant genetics and DNA-damage studies.	Supports annotating rice OsEME1 within a conserved plant replication-stress/HR resolution pathway, though direct rice evidence was sparse before 2025.	Olivier et al., 2016, https://doi.org/10.1105/tpc.15.00898; Herrmann et al., 2015, https://doi.org/10.1093/nar/gkv208 (olivier2016thestructurespecificendonucleases pages 1-5, herrmann2015thenucleasefan1 pages 1-2, olivier2016thestructurespecificendonucleases pages 37-39)
Cross-species inference: 2023 MUS81 complex developments	Recent work shows MUS81 complexes can cleave proteolytically processed DNA-protein crosslink substrates and bulky adduct-containing nicked duplex/3′-flap DNAs, but not native intact TOP1-DNA complexes.	Human/yeast MUS81 complexes cleaved trypsinized DPC substrates; native TOP1 adducts were refractory; MUS81-EME2 and Mus81-Mms4 could be more efficient than MUS81-EME1 in some assays.	2023 biochemical DPC-repair assays in human and yeast MUS81 complexes; nuclear extracts from MUS81-KO vs control cells.	Suggests a broader conserved role for MUS81-family nuclease systems in replication-stress and DPC processing after prior proteolysis/remodeling, relevant as cautious mechanistic inference for plant EME1.	Marini et al., 2023, https://doi.org/10.1186/s12915-023-01614-1; Tomaštíková et al., 2023, https://doi.org/10.1093/plcell/koad020 (marini2023mus81cleavestop1derived pages 2-4, marini2023mus81cleavestop1derived pages 4-6, marini2023mus81cleavestop1derived pages 1-2, tomastikova2023smc56complexmediatedsumoylation pages 2-4)

Table: This table compiles the main experimental evidence for rice OsEME1 (Q0J9J6/LOC_Os04g55500), including substrate specificity, interaction with OsMUS81, critical residues, and mutant phenotypes. It also adds clearly labeled cross-species context from Arabidopsis and 2023 MUS81 literature to support functional annotation where direct rice evidence is limited.

10) Cited visual evidence (figures)

OsEME1 cleavage assays on junction substrates (Y12 / pre-X / X) (du2025essentialmeioticendonuclease media c8ee333c, du2025essentialmeioticendonuclease media 4403c32e)
OsEME1–OsMUS81 interaction and nuclear localization signal in interaction assays (du2025essentialmeioticendonuclease media 8341a419, du2025essentialmeioticendonuclease media 730e3e5a)

11) Reference URLs and publication dates (from retrieved sources)

Du Y. et al. Plant Biotechnology Journal (May 2025): https://doi.org/10.1111/pbi.70101 (du2025essentialmeioticendonuclease pages 1-2)
Mimida N. et al. Plant & Cell Physiology (Apr 2007): https://doi.org/10.1093/pcp/pcm029 (mimida2007twoalternativelyspliced pages 2-3)
Geuting V. et al. Plant Physiology (Apr 2009): https://doi.org/10.1104/pp.109.136846 (geuting2009twodistinctmus81eme1 pages 1-2)
Olivier M. et al. The Plant Cell (Dec 2016): https://doi.org/10.1105/tpc.15.00898 (olivier2016thestructurespecificendonucleases pages 1-5)
Herrmann N.J. et al. Nucleic Acids Research (Mar 2015): https://doi.org/10.1093/nar/gkv208 (herrmann2015thenucleasefan1 pages 1-2)
Tomaštíková E.D. et al. The Plant Cell (Jan 2023): https://doi.org/10.1093/plcell/koad020 (tomastikova2023smc56complexmediatedsumoylation pages 2-4)
Marini V. et al. BMC Biology (May 2023): https://doi.org/10.1186/s12915-023-01614-1 (marini2023mus81cleavestop1derived pages 1-2)

12) Limitations and confidence notes

The strongest rice-specific functional annotation evidence available in the retrieved corpus is from a 2025 study (not 2023–2024). Recent (2023) advances primarily refine MUS81-complex biology and plant DPC repair context but do not directly test rice OsEME1. (du2025essentialmeioticendonuclease pages 12-13, tomastikova2023smc56complexmediatedsumoylation pages 2-4)
Statements about DPC processing and detailed cell-cycle regulation are therefore presented as conserved-pathway inference rather than direct rice OsEME1 experimental outcomes. (marini2023mus81cleavestop1derived pages 4-6, tomastikova2023smc56complexmediatedsumoylation pages 2-4)

References

(du2025essentialmeioticendonuclease pages 2-4): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(du2025essentialmeioticendonuclease pages 11-12): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(olivier2016thestructurespecificendonucleases pages 1-5): Margaux Olivier, Olivier Da Ines, Simon Amiard, Heïdi Serra, Chantal Goubely, Charles I. White, and Maria E. Gallego. The structure-specific endonucleases mus81 and send1 are essential for telomere stability in arabidopsis. Plant Cell, 28:74-86, Dec 2016. URL: https://doi.org/10.1105/tpc.15.00898, doi:10.1105/tpc.15.00898. This article has 21 citations and is from a highest quality peer-reviewed journal.
(geuting2009twodistinctmus81eme1 pages 1-2): Verena Geuting, Daniela Kobbe, Frank Hartung, Jasmin Dürr, Manfred Focke, and Holger Puchta. Two distinct mus81-eme1 complexes from arabidopsis process holliday junctions. Plant Physiology, 150(2):1062-1071, Apr 2009. URL: https://doi.org/10.1104/pp.109.136846, doi:10.1104/pp.109.136846. This article has 64 citations and is from a highest quality peer-reviewed journal.
(geuting2009twodistinctmus81eme1 pages 5-8): Verena Geuting, Daniela Kobbe, Frank Hartung, Jasmin Dürr, Manfred Focke, and Holger Puchta. Two distinct mus81-eme1 complexes from arabidopsis process holliday junctions. Plant Physiology, 150(2):1062-1071, Apr 2009. URL: https://doi.org/10.1104/pp.109.136846, doi:10.1104/pp.109.136846. This article has 64 citations and is from a highest quality peer-reviewed journal.
(du2025essentialmeioticendonuclease pages 12-13): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(du2025essentialmeioticendonuclease pages 7-11): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(du2025essentialmeioticendonuclease media c8ee333c): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(du2025essentialmeioticendonuclease media 8341a419): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(du2025essentialmeioticendonuclease pages 5-7): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(mimida2007twoalternativelyspliced pages 2-3): Naozumi Mimida, Hiroko Kitamoto, Keishi Osakabe, Marina Nakashima, Yuji Ito, Wolf-Dietrich Heyer, Seiichi Toki, and Hiroaki Ichikawa. Two alternatively spliced transcripts generated from osmus81, a rice homolog of yeast mus81, are up-regulated by dna-damaging treatments. Plant & cell physiology, 48 4:648-54, Apr 2007. URL: https://doi.org/10.1093/pcp/pcm029, doi:10.1093/pcp/pcm029. This article has 22 citations and is from a domain leading peer-reviewed journal.
(marini2023mus81cleavestop1derived pages 1-2): Victoria Marini, Fedor Nikulenkov, Pounami Samadder, Sissel Juul, Birgitta R. Knudsen, and Lumir Krejci. Mus81 cleaves top1-derived lesions and other dna–protein cross-links. BMC Biology, May 2023. URL: https://doi.org/10.1186/s12915-023-01614-1, doi:10.1186/s12915-023-01614-1. This article has 16 citations and is from a domain leading peer-reviewed journal.
(marini2023mus81cleavestop1derived pages 4-6): Victoria Marini, Fedor Nikulenkov, Pounami Samadder, Sissel Juul, Birgitta R. Knudsen, and Lumir Krejci. Mus81 cleaves top1-derived lesions and other dna–protein cross-links. BMC Biology, May 2023. URL: https://doi.org/10.1186/s12915-023-01614-1, doi:10.1186/s12915-023-01614-1. This article has 16 citations and is from a domain leading peer-reviewed journal.
(tomastikova2023smc56complexmediatedsumoylation pages 2-4): Eva Dvořák Tomaštíková, Klara Prochazkova, Fen Yang, Jitka Jemelkova, Andreas Finke, Annika Dorn, Mahmoud Said, Holger Puchta, and Ales Pecinka. Smc5/6 complex-mediated sumoylation stimulates dna–protein cross-link repair in arabidopsis. The Plant Cell, 35:1532-1547, Jan 2023. URL: https://doi.org/10.1093/plcell/koad020, doi:10.1093/plcell/koad020. This article has 18 citations.
(geuting2009twodistinctmus81eme1 pages 2-3): Verena Geuting, Daniela Kobbe, Frank Hartung, Jasmin Dürr, Manfred Focke, and Holger Puchta. Two distinct mus81-eme1 complexes from arabidopsis process holliday junctions. Plant Physiology, 150(2):1062-1071, Apr 2009. URL: https://doi.org/10.1104/pp.109.136846, doi:10.1104/pp.109.136846. This article has 64 citations and is from a highest quality peer-reviewed journal.
(verma2020structuralaspectsof pages 11-11): Prabha Verma, Reetika Tandon, Gitanjali Yadav, and Vineet Gaur. Structural aspects of dna repair and recombination in crop improvement. Frontiers in Genetics, Sep 2020. URL: https://doi.org/10.3389/fgene.2020.574549, doi:10.3389/fgene.2020.574549. This article has 30 citations and is from a peer-reviewed journal.
(du2025essentialmeioticendonuclease pages 1-2): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(mimida2007twoalternativelyspliced pages 4-5): Naozumi Mimida, Hiroko Kitamoto, Keishi Osakabe, Marina Nakashima, Yuji Ito, Wolf-Dietrich Heyer, Seiichi Toki, and Hiroaki Ichikawa. Two alternatively spliced transcripts generated from osmus81, a rice homolog of yeast mus81, are up-regulated by dna-damaging treatments. Plant & cell physiology, 48 4:648-54, Apr 2007. URL: https://doi.org/10.1093/pcp/pcm029, doi:10.1093/pcp/pcm029. This article has 22 citations and is from a domain leading peer-reviewed journal.
(herrmann2015thenucleasefan1 pages 1-2): Natalie J. Herrmann, Alexander Knoll, and Holger Puchta. The nuclease fan1 is involved in dna crosslink repair in arabidopsis thaliana independently of the nuclease mus81. Nucleic Acids Research, 43:3653-3666, Mar 2015. URL: https://doi.org/10.1093/nar/gkv208, doi:10.1093/nar/gkv208. This article has 18 citations and is from a highest quality peer-reviewed journal.
(olivier2016thestructurespecificendonucleases pages 37-39): Margaux Olivier, Olivier Da Ines, Simon Amiard, Heïdi Serra, Chantal Goubely, Charles I. White, and Maria E. Gallego. The structure-specific endonucleases mus81 and send1 are essential for telomere stability in arabidopsis. Plant Cell, 28:74-86, Dec 2016. URL: https://doi.org/10.1105/tpc.15.00898, doi:10.1105/tpc.15.00898. This article has 21 citations and is from a highest quality peer-reviewed journal.
(marini2023mus81cleavestop1derived pages 2-4): Victoria Marini, Fedor Nikulenkov, Pounami Samadder, Sissel Juul, Birgitta R. Knudsen, and Lumir Krejci. Mus81 cleaves top1-derived lesions and other dna–protein cross-links. BMC Biology, May 2023. URL: https://doi.org/10.1186/s12915-023-01614-1, doi:10.1186/s12915-023-01614-1. This article has 16 citations and is from a domain leading peer-reviewed journal.
(du2025essentialmeioticendonuclease media 4403c32e): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.
(du2025essentialmeioticendonuclease media 730e3e5a): Yanxin Du, Yang Li, Weijiang Tang, Weiping Mo, Tingting Ma, and Rongcheng Lin. Essential meiotic endonuclease 1 is required for chloroplast development and dna repair in rice. Plant Biotechnology Journal, 23:2931-2948, May 2025. URL: https://doi.org/10.1111/pbi.70101, doi:10.1111/pbi.70101. This article has 0 citations and is from a highest quality peer-reviewed journal.

Artifacts

Edison artifact artifact-00

Citations

olivier2016thestructurespecificendonucleases pages 1-5
du2025essentialmeioticendonuclease pages 11-12
du2025essentialmeioticendonuclease pages 7-11
du2025essentialmeioticendonuclease pages 2-4
du2025essentialmeioticendonuclease pages 5-7
mimida2007twoalternativelyspliced pages 2-3
verma2020structuralaspectsof pages 11-11
du2025essentialmeioticendonuclease pages 1-2
du2025essentialmeioticendonuclease pages 12-13
mimida2007twoalternativelyspliced pages 4-5
olivier2016thestructurespecificendonucleases pages 37-39
https://doi.org/10.1111/pbi.70101
https://doi.org/10.1093/pcp/pcm029
https://doi.org/10.1104/pp.109.136846
https://doi.org/10.1105/tpc.15.00898;
https://doi.org/10.1093/nar/gkv208
https://doi.org/10.1186/s12915-023-01614-1;
https://doi.org/10.1093/plcell/koad020
https://doi.org/10.1105/tpc.15.00898
https://doi.org/10.1186/s12915-023-01614-1
https://doi.org/10.1111/pbi.70101,
https://doi.org/10.1105/tpc.15.00898,
https://doi.org/10.1104/pp.109.136846,
https://doi.org/10.1093/pcp/pcm029,
https://doi.org/10.1186/s12915-023-01614-1,
https://doi.org/10.1093/plcell/koad020,
https://doi.org/10.3389/fgene.2020.574549,
https://doi.org/10.1093/nar/gkv208,

📚 Additional Documentation

Notes

(EME1-notes.md)

EME1 (OsEME1) — Oryza sativa subsp. japonica — research notes

UniProt: Q0J9J6 | Gene: EME1 / OsEME1 | Locus: Os04g0648700, LOC_Os04g55500
"Crossover junction endonuclease EME1" / "Essential meiotic endonuclease 1"; EC 3.1.22.-
Family: EME1/MMS4 family; PANTHER PTHR21077:SF5 (CROSSOVER JUNCTION ENDONUCLEASE MMS4)

1. What EME1/MUS81 is — conserved biology

EME1 (Eme1 in fission yeast; Mms4 in budding yeast; EME1/EME2 in humans) is the
non-catalytic regulatory subunit of a heterodimeric, structure-specific DNA
endonuclease whose catalytic subunit is MUS81. EME1 itself lacks endonuclease
activity in yeast and human; the interaction with MUS81 stimulates and regulates
MUS81's endonucleolytic activity.

PMID:40333587
PMID:40333587

MUS81-EME1 is one of three structure-selective endonuclease systems
(MUS81-EME1/MMS4, SLX1-SLX4, GEN1/Yen1) that resolve branched DNA
intermediates ("Holliday junction resolvases" in the broad sense). MUS81-EME1
has substrate preference for nicked Holliday junctions, 3'-flaps, D-loops and
replication-fork structures — i.e. branched substrates with a 5'-end at the
branch — and resolves intact HJs only with reduced efficiency.

PMID:24008669
PMID:24008669

UniProt FUNCTION (Q0J9J6, by similarity): "Interacts with MUS81 to form a DNA
structure-specific endonuclease with substrate preference for branched DNA
structures with a 5'-end at the branch nick. Typical substrates include 3'-flap
structures, D-loops, replication forks, nicked Holliday junctions and also
intact Holliday junctions with a reduced efficiency. May be required in mitosis
for the processing of stalled or collapsed replication fork intermediates.
Plays a role in DNA repair and in genotoxic stress-induced homologous
recombination (HR) in somatic cells. Mediates a subset of meiotic recombination
events that are insensitive to crossover interference."

This dual mitotic/meiotic role is key: MUS81-EME1 functions in BOTH (a) somatic
(mitotic) replication-fork rescue and DSB repair AND (b) meiotic crossover
formation (the interference-insensitive / "class II" CO pathway). It is NOT a
meiosis-restricted enzyme.

DNA-damage-responsive regulation

MUS81-EME1 activity is cell-cycle and DNA-damage regulated. In fission yeast,
DNA-damage-induced activation of Mus81-Eme1 requires Cdc2(CDK1)- and
Rad3(ATR)-dependent phosphorylation of Eme1; the complex is activated at G2/M.
PMID:23584455
PMID:40333587

In meiosis, MUS81-EME1 (with Yen1/GEN1 and SLX1-SLX4) must be kept inactive
during prophase I to allow proper CO patterning; precocious activation disrupts
crossover distribution.
PMID:29920281

2. OsEME1 — the primary rice paper (Du et al. 2025, Plant Biotechnol J)

PMID:40333587 / DOI:10.1111/pbi.70101 — "ESSENTIAL MEIOTIC ENDONUCLEASE 1 is
required for chloroplast development and DNA repair in rice." This is the
definitive functional characterization of OsEME1 (Q0J9J6).

Key findings:

OsEME1 is a single-copy gene in rice. Unlike Arabidopsis (which has EME1A
and EME1B), rice has only one EME1.
PMID:40333587
OsEME1 is an endonuclease with intrinsic DNA cleavage activity in vitro.
Recombinant MBP-OsEME1 and the C-terminal fragment (OsEME1-C, containing the
ERCC4 domain) both bind and cleave the Y12 branched substrate. This is notable:
in yeast/human, EME1 is the non-catalytic subunit. In the rice in vitro assay,
OsEME1 alone produced cleavage products.
PMID:40333587
PMID:40333587
PMID:40333587
Four conserved residues required for activity. Point mutants of four
conserved residues (modelled on mammalian EME1 catalytic residues) reduced
binding/cleavage, confirming the ERCC4 domain is the catalytic region.
PMID:40333587
OsEME1 binds Mg2+/branched DNA substrates — substrate Kd ~16 µM for Y12.
UniProt cofactor: Mg(2+), Ca(2+).
OsEME1 interacts with OsMUS81. Y2H, luciferase complementation (LCI) and
BiFC all confirm interaction; the ERCC4 domain of OsEME1 and the HhH motif of
OsMUS81 mediate the interaction.
PMID:40333587
PMID:40333587
OsMUS81 binds X-structure substrates but did not itself add cleavage in
the in vitro assay — in the rice system OsEME1-C was the cleaver.
PMID:40333587
Nuclear localization. OsEME1-GFP localizes to the nucleus and overlaps with
the nuclear marker H2B-mCherry.
PMID:40333587
DNA-damage phenotypes. EMS mutant (pale1) and CRISPR oseme1 mutants are
hypersensitive to the DNA-damaging agents MMS and Zeocin (reduced shoot/root
growth). Mutants show increased S/G2 DNA content (cell-cycle arrest), more
S-phase (EdU+) cells, and increased γH2AX foci (a DSB / DNA-damage marker)
with or without Zeocin.
PMID:40333587
PMID:40333587
OsEME1 is involved in HR repair. The proposed model: under high light or
genotoxic stress, DSBs form, the DNA-damage response is activated, repair
proceeds via HR, double Holliday junctions accumulate, and OsEME1 is recruited
to bind and cleave the HJ substrates, maintaining genome stability.
PMID:40333587
Chloroplast / striped-leaf phenotype. oseme1 mutants have a striped-albino
leaf phenotype, defective chloroplast development, reduced chlorophyll/
carotenoids, low PSII efficiency; the phenotype is strongly enhanced by
high-light stress. OsEME1 globally regulates expression of photosynthesis and
DNA-repair genes. Mechanistic model: OsEME1 maintains genome/transcription
integrity of nuclear chloroplast-development genes (e.g. OsGLK1/OsGLK2), which
in turn drive chloroplast development. The chloroplast phenotype is a
downstream consequence of genome-maintenance failure, not a direct chloroplast
function (OsEME1 is nuclear).
PMID:40333587
PMID:40333587
High-light sensitivity is real and experimentally demonstrated in this
paper (oseme1 mutants are hypersensitive to high light, with severe leaf
striping and cell death after HL treatment). This is genuine functional
evidence — but note this paper is PMID:40333587, NOT the IEP reference
PMID:12869764.

3. Rice OsMUS81 — meiosis (Mu et al. 2022, New Phytol)

PMID:36495065 / DOI:10.1111/nph.18668 — "MUS81 is required for atypical
recombination intermediate resolution but not crossover designation in rice."

Rice mus81 mutants have normal chiasma numbers and normal HEI10 foci (the
interference-sensitive / class I CO pathway). MUS81 does NOT contribute
substantially to crossover designation in rice.
PMID:36495065
PMID:36495065
Instead, MUS81 resolves atypical / aberrant meiotic recombination
intermediates — mus81 zep1 and mus81 fancm double mutants show chromosome
fragments and bridges.
PMID:36495065
PMID:36495065
FANCM-dependent extra (interference-insensitive) COs in rice fancm mutants
require MUS81 for resolution.
PMID:37632767

So the rice MUS81-EME1 complex has a meiotic role — but it is a minor/atypical-
intermediate-resolution role, NOT a meiosis-restricted "essential meiotic
endonuclease" role. The "Essential meiotic endonuclease" name is historical
(from S. pombe Eme1) and is somewhat misleading for the plant ortholog.

4. Other plant context

OsGEN1 (rice GEN1/Yen1 ortholog): a canonical HJ resolvase of the RAD2/XPG
family; works alongside MUS81-EME1 and SLX1-SLX4 in HR-mediated DNA repair.
osgen1 mutants show ~6% reduced chiasma frequency and persistent DSBs in
microspores leading to male sterility.
PMID:28049740
Arabidopsis: AtMUS81 with AtEME1A/AtEME1B. MUS81 defines a DNA-repair pathway
parallel to RECQ4A; recq4A mus81 double mutants are lethal (rescued by
blocking HR), showing MUS81 processes recombination-induced intermediates
during replication. MUS81 is required for efficient synthesis-dependent strand
annealing (SDSA).
PMID:20971895
PMID:20971895
PMID:24635147
AtEME1A/AtEME1B are recruited to Holliday junction substrates and cleave at
nicks in the DNA strands.
PMID:40333587

5. Assessment of the IEP annotations from PMID:12869764

PMID:12869764 (Kikuchi et al. 2003, Science) is "Collection, mapping, and
annotation of over 28,000 cDNA clones from japonica rice." It is a large-scale
full-length cDNA sequencing/annotation consortium paper. Its abstract describes
ONLY cDNA collection, sequencing, mapping to genomic DNA, and InterPro-based
function assignment. There is NO gene-specific expression-profiling experiment
for EME1 and NO experiment testing response to UV, gamma radiation or high
light.

The three IEP annotations (response to UV GO:0009411, response to high light
intensity GO:0009644, response to gamma radiation GO:0010332) all cite
PMID:12869764 as the IEP (Inferred from Expression Pattern) reference. This is
almost certainly a curation artefact: the rice FL-cDNA consortium built cDNA
libraries from many tissues and stress conditions, and a clone for this gene may
have come from a stress-treated library (UV-/gamma-/high-light-treated tissue).
The presence of a cDNA clone in a stress library is library-of-origin
information, NOT an expression-response measurement, and certainly not evidence
of biological "involvement in" a stress response. IEP from this kind of source
is not appropriate functional evidence. Expression ≠ function.

Note: high-light sensitivity of OsEME1 IS genuinely demonstrated — but in
PMID:40333587 (oseme1 mutants hypersensitive to high light), as a downstream
consequence of a genome-maintenance defect, not a primary "response to high
light" function. The IEP annotations remain unsupported by their cited
reference.

6. Summary of SPKW-retired annotation assessment

The six retired SPKW (GO_REF:0000043) annotations were keyword-to-GO mappings:
- meiotic cell cycle (GO:0051321) <- KW "Meiosis" — OVER-SPECIFIC. EME1/MUS81 is
not meiosis-restricted; its dominant role in rice is somatic DSB/replication
repair. Removal justified.
- cell division (GO:0051301) <- KW "Cell division" — weakly supported, generic;
EME1 acts in cell-cycle-coupled repair but "cell division" per se is not its
function. Removal justified (over-annotation).
- DNA recombination (GO:0006310) <- KW "DNA recombination" — correct in essence
(EME1 resolves HR intermediates) but generic; better captured by HR-repair /
resolution terms. Removal acceptable; correct biology retained by current
IBA/IEA terms.
- endonuclease activity (GO:0004519) <- KW "Endonuclease/Nuclease" — GENUINELY
CORRECT. EME1 IS an endonuclease (EC 3.1.22.-), demonstrated biochemically in
rice (PMID:40333587). Current GOA has NO catalytic MF term — only "DNA
binding". Removing endonuclease activity LOST a correct, core annotation. This
removal was NOT justified; a NEW catalytic MF term should be added.
- hydrolase activity (GO:0016787) <- KW "Hydrolase" — correct but vague parent
of endonuclease activity. Removal of the vague term is acceptable IF a
specific endonuclease MF is present (it is not — see above).
- metal ion binding (GO:0046872) <- KW "Metal-binding" — correct (Mg2+/Ca2+
cofactor) but the keyword mapping is generic. MUS81-EME1 nuclease chemistry is
Mg2+-dependent. Removal of the generic term is acceptable; magnesium ion
binding (GO:0000287) would be the accurate specific term if retained.

📄 View Raw YAML

id: Q0J9J6
gene_symbol: EME1
product_type: PROTEIN
status: COMPLETE
taxon:
  id: NCBITaxon:39947
  label: Oryza sativa subsp. japonica
description: >
  OsEME1 (Q0J9J6) is the rice ortholog of the EME1/MMS4-family DNA structure-specific
  endonuclease subunit. EME1 partners with MUS81 to form a heterodimeric (heterotetrameric)
  endonuclease that binds and cleaves branched DNA intermediates - nicked Holliday
  junctions, 3'-flaps, D-loops and replication-fork structures - generated during
  homologous recombination (HR) repair and replication. Unlike yeast Mms4/Eme1 and
  human EME1, which are non-catalytic regulatory subunits, recombinant OsEME1 (via its
  C-terminal ERCC4 domain) directly binds and cleaves branched DNA substrates in vitro,
  with four conserved residues required for activity (Du et al. 2025, PMID:40333587).
  OsEME1 is a single-copy nuclear gene; the OsEME1-GFP fusion localizes to the nucleus.
  Its dominant biological role in rice is somatic DNA-damage repair: oseme1 loss-of-function
  mutants are hypersensitive to the DNA-damaging agents MMS and Zeocin, accumulate
  gamma-H2AX foci, show S/G2 cell-cycle arrest, and have a striped-albino-leaf /
  defective-chloroplast phenotype that is strongly enhanced by high light. The chloroplast
  defect is a downstream consequence of nuclear genome-maintenance failure (OsEME1
  cleaves damaged DNA in nuclear chloroplast-development genes such as OsGLK1/OsGLK2).
  Consistent with the rice MUS81 meiosis study (Mu et al. 2022, PMID:36495065), the
  MUS81-EME1 complex in rice has only a minor meiotic role - resolution of atypical
  meiotic recombination intermediates - and is NOT required for crossover designation;
  the "Essential meiotic endonuclease" name is historical (from S. pombe eme1) and is
  misleading for the plant protein, which acts predominantly in mitotic/somatic HR
  repair and replication-fork rescue.
existing_annotations:
# --- SPKW keyword-mapping annotations (GO_REF:0000043) ---
# Present in the Sept 2025 goa_uniprot_gcrp snapshot (go-db plant.ddb); REMOVED
# from the current (2026) GOA release when GOA retired the keyword2GO pipeline
# for cellular organisms. Reviewed retrospectively to assess whether removal was
# justified. These six are the TRUE SPKW-unique set (closure-filtered).
- term:
    id: GO:0051321
    label: meiotic cell cycle
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  retired: true
  review:
    summary: >
      SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "Meiosis";
      snapshot-only, removed in the current GOA release. EME1/MUS81 is a structure-specific
      endonuclease acting on branched DNA in BOTH mitotic/somatic DNA repair and meiotic
      recombination - it is not meiosis-restricted.
    action: MARK_AS_OVER_ANNOTATED
    reason: >
      GOA's removal of this annotation was JUSTIFIED. The keyword "Meiosis" reflects the
      historical "Essential meiotic endonuclease" name (inherited from S. pombe eme1) but
      mapping it to "meiotic cell cycle" is over-specific for the rice protein. The dominant,
      experimentally demonstrated role of OsEME1 in rice is somatic DNA-damage repair:
      oseme1 mutants are hypersensitive to MMS/Zeocin, accumulate gamma-H2AX foci and show
      cell-cycle arrest in vegetative tissue, with a striped-leaf/chloroplast phenotype
      [PMID:40333587]. The rice MUS81-EME1 complex contributes only minimally to meiosis -
      mus81 mutants have normal chiasma numbers and normal crossover designation and the
      complex acts only on atypical meiotic intermediates [PMID:36495065]. A keyword-based
      blanket "meiotic cell cycle" term is therefore an over-annotation; the genuine meiotic
      contribution is more precisely captured by the IBA term "resolution of meiotic
      recombination intermediates" (GO:0000712), which is retained in current GOA.
    supported_by:
    - reference_id: PMID:36495065
      supporting_text: "the total chiasma numbers in mus81 mutants were indistinguishable
        from wild-type. The numbers of HEI10 foci ... in mus81 were also similar to that
        of wild-type."
    - reference_id: PMID:40333587
      supporting_text: "treatment with 75 μg/mL of MMS or 50 μg/mL of Zeocin
        strongly inhibited root and shoot growth, especially in the mutants"
    - reference_id: PMID:40333587
      supporting_text: "OsEME1 regulates chloroplast development and division by maintaining
        the transcription of chloroplast-related genes and nuclear genome integrity"
- term:
    id: GO:0051301
    label: cell division
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  retired: true
  review:
    summary: >
      SPKW (GO_REF:0000043) annotation derived from the UniProt keywords "Cell division"
      / "Mitosis"; snapshot-only, removed in current GOA. EME1 acts in cell-cycle-coupled
      DNA repair, but "cell division" per se is not its molecular role.
    action: MARK_AS_OVER_ANNOTATED
    reason: >
      GOA's removal of this annotation was JUSTIFIED. The keyword mapping produces an overly
      generic process term. OsEME1 is a DNA-repair endonuclease whose activity is cell-cycle
      regulated (MUS81-EME1 is activated at G2/M and during the DNA-damage response in other
      species [PMID:23584455]), and oseme1 mutants show cell-cycle arrest as a CONSEQUENCE
      of unrepaired DNA damage [PMID:40333587]. The protein does not have a direct role in
      the cell-division process itself (e.g. cytokinesis, chromosome segregation machinery).
      "Cell division" is an indirect downstream/contextual term and is better replaced by
      specific DNA-repair process terms (DNA repair, double-strand break repair) that are
      retained in current GOA.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "The DNA contents at the S and G2 stages following Zeocin treatment
        increased in ... cells compared to KY131"
    - reference_id: PMID:40333587
      supporting_text: "mutations in OsEME1 led to cell cycle arrest and a DNA damage response"
- term:
    id: GO:0006310
    label: DNA recombination
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  retired: true
  review:
    summary: >
      SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "DNA recombination";
      snapshot-only, removed in current GOA. EME1/MUS81 resolves homologous-recombination
      intermediates, so the essence of the term is biologically correct, but the keyword
      mapping produces a non-specific parent term.
    action: MARK_AS_OVER_ANNOTATED
    reason: >
      GOA's removal of this annotation was reasonable and did NOT lose correct biology.
      OsEME1 genuinely participates in homologous recombination - it binds and cleaves
      branched HR intermediates (Y12, pre-X12, X12 substrates) in vitro [PMID:40333587] -
      so "DNA recombination" is not wrong. However, it is a broad parent term, and the
      specific HR-repair function is better and still represented in current GOA by the
      IBA terms "double-strand break repair" (GO:0006302) and "resolution of meiotic
      recombination intermediates" (GO:0000712) and the IEA term "DNA repair" (GO:0006281).
      A NEW more precise term, "double-strand break repair via homologous recombination"
      (GO:0000724), is proposed below. Removal of the generic keyword-derived term is
      therefore acceptable.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which
        are typical substrates after HR repair of DNA damage"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "rice OsEME1 likely participates in the canonical SSE pathway for
        processing recombination/replication structures"
- term:
    id: GO:0004519
    label: endonuclease activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  retired: true
  review:
    summary: >
      SPKW (GO_REF:0000043) annotation derived from the UniProt keywords
      "Endonuclease"/"Nuclease"; snapshot-only, removed in current GOA. EME1 IS a
      structure-specific endonuclease (EC 3.1.22.-), and recombinant OsEME1 directly binds
      and cleaves branched DNA substrates in vitro via its C-terminal ERCC4 domain.
    action: MODIFY
    reason: >
      GOA's removal of this annotation was NOT justified - a correct, core molecular-function
      annotation was lost. EME1 is genuinely an endonuclease: the UniProt RecName is
      "Crossover junction endonuclease EME1" with EC 3.1.22.-, and Du et al. (2025) showed
      directly that recombinant MBP-OsEME1 and its C-terminal fragment bind and cleave the
      branched Y12/pre-X12/X12 substrates, with four conserved residues required for activity
      [PMID:40333587]. Critically, the current (2026) GOA release contains NO catalytic
      molecular-function term for OsEME1 at all - only "DNA binding" (GO:0003677). Removing
      "endonuclease activity" without replacement leaves the gene with no MF describing its
      core enzymatic activity. The annotation should be retained, but MODIFIED to the more
      specific and informative term "crossover junction DNA endonuclease activity"
      (GO:0008821), which precisely captures the EME1/MUS81 structure-specific
      branched-DNA cleavage activity (see also the NEW entry below).
    proposed_replacement_terms:
    - id: GO:0008821
      label: crossover junction DNA endonuclease activity
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "The major cleavage products were obtained in OsEME1 and OsEME1-C,
        and their levels gradually increased over time"
    - reference_id: PMID:40333587
      supporting_text: "OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which
        are typical substrates after HR repair of DNA damage"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "Purified OsEME1 directly binds and cleaves branched DNA substrates
        typical of HR intermediates, including Y12, pre-X12, and X12 ... Supports
        annotation as a crossover-junction/branched-DNA endonuclease involved in HR-mediated
        DSB repair."
- term:
    id: GO:0016787
    label: hydrolase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  retired: true
  review:
    summary: >
      SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "Hydrolase";
      snapshot-only, removed in current GOA. "Hydrolase activity" is the high-level parent
      of the gene's true endonuclease (phosphodiester-bond hydrolase) activity.
    action: MARK_AS_OVER_ANNOTATED
    reason: >
      GOA's removal of this annotation was JUSTIFIED. "Hydrolase activity" is a very broad
      grouping term; an endonuclease is a hydrolase, so the term is not wrong, but it is
      uninformative. The correct, specific catalytic function (endonuclease / crossover
      junction DNA endonuclease activity) is demonstrated experimentally [PMID:40333587]
      and should be annotated directly (see the MODIFY on GO:0004519 and the NEW
      GO:0008821 entry). Retaining a top-level "hydrolase activity" term provides no
      additional information once the specific endonuclease MF is present. Removal of the
      vague parent is appropriate.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "we measured the endonuclease activity of OsEME1 by performing a
        nuclease assay, followed by gel electrophoresis"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "the most direct biochemical description is that **OsEME1 binds and
        cleaves branched DNA substrates** representing recombination/repair intermediates ...
        This is an **endonucleolytic cleavage** of DNA phosphodiester bonds (EC class **3.1.22.-**)
        on structured substrates."
- term:
    id: GO:0046872
    label: metal ion binding
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  retired: true
  review:
    summary: >
      SPKW (GO_REF:0000043) annotation derived from the UniProt keywords
      "Metal-binding"/"Magnesium"/"Calcium"; snapshot-only, removed in current GOA. The
      MUS81-EME1 nuclease reaction is metal-dependent (Mg2+/Ca2+ cofactors).
    action: MARK_AS_OVER_ANNOTATED
    reason: >
      GOA's removal of this generic keyword-derived term was acceptable. The UniProt entry
      lists Mg(2+) and Ca(2+) as cofactors (by similarity), and structure-specific nuclease
      chemistry is divalent-metal-dependent, so the gene does bind metal ions. However,
      "metal ion binding" is a broad term inferred from a keyword rather than from
      gene-specific experimental evidence, and no metal-coordinating residues are annotated
      on the rice protein. If a metal-binding MF were retained it should be the specific
      "magnesium ion binding" (GO:0000287); but as a free-standing keyword-derived
      annotation the broad term adds little, so its removal is reasonable. The metal
      dependence is implicit in the endonuclease MF.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "Eme1 [methyl methanesulfonate 4 (Mms4) in ...] forms a heterodimeric
        endonuclease with Mus81 ..., cleaving branched DNA substrates such as HJs, 3′-DNA
        flaps and replication forks"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "activity is influenced by divalent cations (Mg2+/Ca2+ support
        activity; Mn2+ can increase activity and product diversity)."
# --- Current GOA annotations (2026 release) ---
- term:
    id: GO:0031297
    label: replication fork processing
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >
      IBA annotation propagated across the EME1/MUS81 phylogenetic group. The MUS81-EME1
      complex processes stalled/collapsed replication-fork intermediates - a conserved
      mitotic function of the complex.
    action: ACCEPT
    reason: >
      Well supported by conserved biology and consistent with the UniProt FUNCTION
      statement ("May be required in mitosis for the processing of stalled or collapsed
      replication fork intermediates"). In Arabidopsis, MUS81 and RECQ4A process
      recombination-induced aberrant intermediates during replication, and recq4A mus81
      double mutants are lethal [PMID:20971895]. Branched fork structures (3'-flaps,
      replication forks) are canonical MUS81-EME1 substrates and OsEME1 cleaves
      replication-fork-type Y/X branched substrates in vitro [PMID:40333587]. The IBA
      annotation is at an appropriate level of specificity.
    supported_by:
    - reference_id: PMID:20971895
      supporting_text: "RECQ4A and MUS81 are required for processing recombination-induced
        aberrant intermediates during replication"
    - reference_id: PMID:24008669
      supporting_text: "RuvC is also known to cut branched DNA intermediates that originate
        directly from blocked replication forks, targeting them for origin-independent
        replication restart"
- term:
    id: GO:0048476
    label: Holliday junction resolvase complex
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >
      IBA annotation: EME1 is a subunit of the MUS81-EME1 structure-specific endonuclease,
      which acts as a Holliday-junction-resolving complex. Confirmed in rice by direct
      OsEME1-OsMUS81 interaction.
    action: ACCEPT
    reason: >
      Strongly supported. EME1/MUS81 heterodimers (heterotetramers) are conserved Holliday-
      junction resolvases across yeast, mammals and plants. In rice, OsEME1 directly
      interacts with OsMUS81 - shown by yeast two-hybrid, luciferase complementation and
      BiFC, with the OsEME1 ERCC4 domain and the OsMUS81 HhH motif mediating the interaction
      [PMID:40333587]. This is a core cellular-component annotation for EME1.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "OsEME1 interacts with OsMUS81 and ... the ERCC4 domain of OsEME1 is
        required for this interaction"
    - reference_id: PMID:40333587
      supporting_text: "Homologues of EME1 and MUS81 interact to form heterotetramers in
        mammals, yeasts and Arabidopsis"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "OsEME1 physically interacts with OsMUS81; interaction depends on the
        OsEME1 ERCC4 domain and OsMUS81 HhH motif, and interaction signal localizes to
        nuclei ... Strong evidence that rice OsEME1 functions as the MUS81 partner in a
        nuclear structure-specific nuclease complex."
- term:
    id: GO:0000712
    label: resolution of meiotic recombination intermediates
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >
      IBA annotation: the MUS81-EME1 complex resolves meiotic recombination intermediates,
      contributing to a subset of (interference-insensitive) crossovers. In rice this role
      is minor and limited to atypical meiotic intermediates.
    action: KEEP_AS_NON_CORE
    reason: >
      The annotation is correct but reflects a minor, non-core function for the rice
      protein. Rice MUS81 is required for resolution of ATYPICAL meiotic recombination
      intermediates (mus81 zep1 and mus81 fancm double mutants show chromosome fragments
      and bridges) and the FANCM-dependent interference-insensitive extra crossovers
      require MUS81 for resolution [PMID:36495065, PMID:37632767]. However, rice mus81
      single mutants have wild-type chiasma numbers and normal crossover designation, so
      meiotic crossover resolution is not the dominant function [PMID:36495065]. Given
      that the demonstrated dominant role of OsEME1 in rice is somatic DSB/replication
      repair [PMID:40333587], this meiotic-intermediate-resolution term should be retained
      but classified as non-core.
    supported_by:
    - reference_id: PMID:36495065
      supporting_text: "MUS81 ... plays a crucial role in the resolution of atypical meiotic
        intermediates by working together with other anti-crossover factors"
    - reference_id: PMID:37632767
      supporting_text: "the meiotic extra COs are not marked with HEI10 and require MUS81
        resolvase for resolution"
- term:
    id: GO:0006302
    label: double-strand break repair
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >
      IBA annotation: EME1/MUS81 functions in double-strand break repair. This is the core
      biological process of OsEME1, supported directly by rice mutant phenotypes.
    action: ACCEPT
    reason: >
      This is a core function and is well supported by direct experimental evidence in rice.
      oseme1 loss-of-function mutants are hypersensitive to the DNA-damaging agents MMS and
      Zeocin, accumulate gamma-H2AX foci (a DSB marker) and show S/G2 cell-cycle arrest,
      and OsEME1 binds and cleaves branched HR-repair intermediates in vitro [PMID:40333587].
      In Arabidopsis, MUS81 is required for efficient synthesis-dependent strand annealing
      (an HR-mediated DSB repair pathway) [PMID:20971895]. The IBA term is at the right
      level of specificity; a more precise term (GO:0000724, double-strand break repair
      via homologous recombination) is proposed below.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "The ... and ... mutants exhibited significantly more γ2HAX
        foci than KY131 with or without Zeocin treatment ... OsEME1 regulates DSB repair
        and the cell cycle"
    - reference_id: PMID:20971895
      supporting_text: "MUS81 and RECQ4A are required for efficient synthesis-dependent
        strand annealing (SDSA)"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "Loss of OsEME1 caused **hypersensitivity to DNA-damaging agents** ...
        consistent with a function in HR-mediated repair and/or recovery from
        replication-associated lesions ... Supports a role in HR-mediated DSB repair and
        replication-associated genome stability."
- term:
    id: GO:0031573
    label: mitotic intra-S DNA damage checkpoint signaling
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >
      IBA annotation propagated from fission yeast, where Mus81-Eme1 activity is integrated
      with the intra-S/DNA-damage checkpoint via ATR(Rad3)/CDK phosphorylation of Eme1.
    action: KEEP_AS_NON_CORE
    reason: >
      The annotation captures a genuine, conserved regulatory connection but is not a core
      function and is supported only indirectly for rice. In S. pombe, DNA-damage-induced
      activation of Mus81-Eme1 requires Cdc2(CDK1)- and Rad3(ATR)-dependent phosphorylation
      of Eme1 [PMID:23584455]. In rice, oseme1 mutants show DNA-damage-induced cell-cycle
      arrest and a DNA-damage response [PMID:40333587], consistent with checkpoint
      coupling, but there is no direct evidence that OsEME1 itself transduces checkpoint
      signaling - it is a downstream effector (an endonuclease) whose activity is checkpoint-
      regulated rather than a checkpoint signaling component. Retain as non-core; OsEME1's
      relationship to the checkpoint is being regulated BY it, not enacting it.
    supported_by:
    - reference_id: PMID:23584455
      supporting_text: "DNA damage-induced activation of Mus81-Eme1 ... requires both
        Cdc2(CDK1)- and Rad3(ATR)-dependent phosphorylation of Eme1"
    - reference_id: PMID:40333587
      supporting_text: "mutations in ... led to cell cycle arrest and a DNA damage response"
- term:
    id: GO:0003677
    label: DNA binding
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  review:
    summary: >
      IEA annotation from InterPro (IPR006166, ERCC4 domain). OsEME1 binds DNA directly -
      confirmed biochemically.
    action: ACCEPT
    reason: >
      Correct. The ERCC4 domain has nucleic-acid-binding activity, and recombinant OsEME1
      and OsEME1-C show major binding shifts on branched Y12 substrates in EMSA, with a
      measured substrate Kd of ~16 microM by microscale thermophoresis [PMID:40333587].
      "DNA binding" is a generic term; the more informative activity is the structure-
      specific endonuclease MF (see the NEW GO:0008821 entry), but the DNA-binding IEA is
      not incorrect and can be accepted.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "Major binding shifts were detected in the OsEME1 and OsEME1-C lines
        but not in the MBP control"
- term:
    id: GO:0005634
    label: nucleus
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  review:
    summary: >
      IEA annotation for nuclear localization. Directly confirmed in rice by GFP imaging.
    action: ACCEPT
    reason: >
      Strongly supported by direct experimental evidence: the OsEME1-GFP fusion protein
      localizes to the nucleus and overlaps with the nuclear marker H2B-mCherry in a
      transient expression assay [PMID:40333587]. Nuclear localization is consistent with
      its role as a nuclear DNA-repair endonuclease and with the UniProt subcellular
      location.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "the OsEME1-GFP fusion protein localized to the nucleus and overlapped
        closely with the nuclear protein H2B-mCherry"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "Direct evidence from interaction imaging assays indicates that the
        **OsEME1–OsMUS81 interaction occurs in the nucleus**, supporting a primary nuclear
        role in processing DNA repair/recombination intermediates."
- term:
    id: GO:0006281
    label: DNA repair
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  review:
    summary: >
      IEA annotation from InterPro (IPR033310, Mms4/EME1/EME2). DNA repair is a core
      biological process of OsEME1.
    action: ACCEPT
    reason: >
      Core function, strongly supported by direct rice evidence. oseme1 mutants are
      hypersensitive to DNA-damaging agents (MMS, Zeocin), accumulate gamma-H2AX foci and
      show DNA-damage-induced cell-cycle arrest [PMID:40333587]. OsEME1 cleaves branched
      HR-repair intermediates in vitro. The term is appropriate, though the more specific
      "double-strand break repair" (GO:0006302, IBA, retained) and the proposed GO:0000724
      better capture the HR-specific mechanism.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "we discovered a biochemical role for OsEME1 in HR repair and a
        biological role for this protein in chloroplast development"
- term:
    id: GO:0048476
    label: Holliday junction resolvase complex
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  review:
    summary: >
      IEA annotation from InterPro (IPR033310) for the Holliday junction resolvase complex;
      duplicates the IBA annotation to the same term.
    action: ACCEPT
    reason: >
      Correct and consistent with the IBA annotation to the same term and with direct
      evidence that OsEME1 interacts with OsMUS81 to form a structure-specific endonuclease
      complex [PMID:40333587]. Duplicate annotations with different evidence codes are
      acceptable; the IEA provides additional computational support for a well-established
      complex membership.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "OsEME1 interacts with OsMUS81 and ... the ERCC4 domain of OsEME1 is
        required for this interaction"
- term:
    id: GO:0009411
    label: response to UV
  evidence_type: IEP
  original_reference_id: PMID:12869764
  review:
    summary: >
      IEP annotation citing PMID:12869764, which is the rice full-length cDNA consortium
      paper ("Collection, mapping, and annotation of over 28,000 cDNA clones from japonica
      rice"). That paper contains no gene-specific UV-response expression experiment.
    action: REMOVE
    reason: >
      This annotation is not supported by its cited reference. PMID:12869764 is a large-scale
      cDNA collection/sequencing/annotation paper; it reports only clone collection, genome
      mapping and InterPro-based function assignment, with no expression-response assay for
      this gene. The IEP (Inferred from Expression Pattern) almost certainly derives from
      the stress-treated source of a cDNA library (library-of-origin metadata), which is not
      a valid measurement of UV-induced expression and does not demonstrate biological
      "involvement in" a UV response. Expression presence is not function. While OsEME1 IS a
      DNA-damage-repair gene (and could plausibly act in UV-DSB repair), the only
      experimental DNA-damage data [PMID:40333587] used MMS and Zeocin, not UV. This
      keyword/library-derived IEP should be removed.
    supported_by:
    - reference_id: PMID:12869764
      supporting_text: "We collected and completely sequenced 28,469 full-length
        complementary DNA clones from Oryza sativa L. ssp. japonica cv. Nipponbare."
- term:
    id: GO:0009644
    label: response to high light intensity
  evidence_type: IEP
  original_reference_id: PMID:12869764
  review:
    summary: >
      IEP annotation citing PMID:12869764 (rice full-length cDNA collection paper), which
      contains no gene-specific high-light-response expression experiment for this gene.
    action: REMOVE
    reason: >
      The cited reference does not support this annotation. PMID:12869764 is a cDNA
      collection/annotation paper with no expression-response assay for OsEME1; the IEP is
      a library-of-origin / keyword artefact, and expression is not function. Notably,
      OsEME1 high-light sensitivity IS genuine - but it was demonstrated in a different,
      later paper (Du et al. 2025, PMID:40333587), where oseme1 mutants show severe leaf
      striping and cell death after high-light treatment. Even there, high-light sensitivity
      is a downstream consequence of a nuclear genome-maintenance defect (failure to repair
      oxidative/replication DNA damage), not a primary "response to high light intensity"
      function of the protein. This particular IEP annotation, as attributed to
      PMID:12869764, is unsupported and should be removed.
    supported_by:
    - reference_id: PMID:12869764
      supporting_text: "Through homology searches of publicly available sequence data, we
        assigned tentative protein functions to 21,596 clones (75.86%)."
- term:
    id: GO:0010332
    label: response to gamma radiation
  evidence_type: IEP
  original_reference_id: PMID:12869764
  review:
    summary: >
      IEP annotation citing PMID:12869764 (rice full-length cDNA collection paper), which
      contains no gene-specific gamma-radiation-response expression experiment.
    action: REMOVE
    reason: >
      Not supported by the cited reference. PMID:12869764 reports cDNA clone collection,
      mapping and annotation, with no gamma-irradiation expression experiment for OsEME1.
      The IEP reflects library-of-origin metadata rather than a measured expression
      response, and expression presence is not evidence of biological involvement. The
      gene's DNA-repair role is real and captured by the DNA-repair / DSB-repair process
      terms, but this specific "response to gamma radiation" IEP is an over-annotation
      attributed to a paper that does not test it, and should be removed.
    supported_by:
    - reference_id: PMID:12869764
      supporting_text: "Mapping of the cDNA clones to genomic DNA revealed that there are
        19,000 to 20,500 transcription units in the rice genome."
# --- NEW annotations proposed from the literature ---
- term:
    id: GO:0008821
    label: crossover junction DNA endonuclease activity
  evidence_type: IDA
  original_reference_id: PMID:40333587
  review:
    summary: >
      OsEME1 is a structure-specific endonuclease that binds and cleaves branched
      Holliday-junction-type DNA substrates. This catalytic molecular function is not
      represented in current GOA and should be added.
    action: NEW
    reason: >
      The current GOA release has NO catalytic MF term for OsEME1 (only "DNA binding"),
      and the retired SPKW "endonuclease activity" was the only catalytic annotation lost.
      Du et al. (2025) demonstrated directly that recombinant OsEME1 / OsEME1-C bind and
      cleave the branched Y12, pre-X12 and X12 substrates (typical Holliday-junction /
      crossover-junction structures generated during HR), and that four conserved ERCC4-domain
      residues are required for activity [PMID:40333587]. The EME1/MUS81 complex is the
      canonical crossover-junction (Holliday junction) resolvase. "Crossover junction DNA
      endonuclease activity" (GO:0008821) is the accurate, specific MF and corresponds to
      the UniProt RecName "Crossover junction endonuclease EME1" and EC 3.1.22.-. IDA is
      justified by the in vitro nuclease/EMSA assays.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which
        are typical substrates after HR repair of DNA damage"
    - reference_id: PMID:40333587
      supporting_text: "these four conserved amino acids are essential for the endonuclease
        activity of OsEME1"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "Conserved residues L439, P487, E490, and K494 are important for
        activity; L439V, E490A, and K494E markedly reduce binding/cleavage ... Identifies
        functionally critical conserved residues for annotation of catalytic
        mechanism/structure-function relationships."
- term:
    id: GO:0000724
    label: double-strand break repair via homologous recombination
  evidence_type: IMP
  original_reference_id: PMID:40333587
  review:
    summary: >
      OsEME1 acts in homologous-recombination-mediated repair of DNA double-strand breaks,
      resolving the branched HR intermediates (Holliday junctions) that arise during repair.
    action: NEW
    reason: >
      Current GOA captures "double-strand break repair" (GO:0006302, IBA) and "DNA repair"
      (GO:0006281, IEA) but not the specific HR mechanism. Du et al. (2025) show that
      oseme1 mutants are hypersensitive to DNA-damaging agents, accumulate gamma-H2AX foci
      and show DNA-damage-induced cell-cycle arrest, and that OsEME1 binds/cleaves the
      branched substrates produced after HR repair; the authors explicitly conclude a
      "biochemical role for OsEME1 in HR repair" [PMID:40333587]. In Arabidopsis, MUS81 is
      required for synthesis-dependent strand annealing, an HR sub-pathway [PMID:20971895].
      GO:0000724 is the precise process term. IMP is justified by the oseme1 loss-of-function
      DNA-damage-sensitivity phenotypes.
    supported_by:
    - reference_id: PMID:40333587
      supporting_text: "This study highlights the role of OsEME1 in regulating chloroplast
        development by modulating homologous recombination repair in response to damage to
        double-stranded DNA"
    - reference_id: PMID:40333587
      supporting_text: "The MUS81–EME complex directly binds to and cleaves typical DNA
        substrates produced after HR repair, a process conserved in yeasts, mammals, dicots
        and monocots"
    - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
      supporting_text: "OsEME1 is best annotated as a **nuclear, structure-specific
        endonuclease** that cleaves **branched/junction DNA substrates** consistent with
        HR/replication intermediates ... rice OsEME1 likely participates in the canonical
        SSE pathway for processing recombination/replication structures"
references:
- id: GO_REF:0000002
  title: Gene Ontology annotation through association of InterPro records with GO
    terms
  findings:
  - statement: InterPro-to-GO mappings (IPR006166 ERCC4 domain; IPR033310 Mms4/EME1/EME2)
      assign DNA binding, DNA repair and Holliday junction resolvase complex to EME1.
- id: GO_REF:0000043
  title: Gene Ontology annotation based on UniProtKB/Swiss-Prot keyword mapping
  findings:
  - statement: SwissProt keyword-derived (SPKW) annotations present in the Sept 2025
      goa_uniprot_gcrp snapshot but removed from the current GOA release after GOA
      retired the keyword2GO pipeline for cellular organisms.
  - statement: For EME1, the keyword "Endonuclease" mapped to a genuinely correct catalytic
      MF; its removal without replacement left the gene with no catalytic function term.
- id: GO_REF:0000033
  title: Annotation inferences using phylogenetic trees
  findings:
  - statement: EME1/MUS81 functions (Holliday junction resolvase complex, replication fork
      processing, double-strand break repair, meiotic recombination intermediate resolution)
      are conserved across the PTHR21077 phylogenetic group.
- id: GO_REF:0000120
  title: Combined Automated Annotation using Multiple IEA Methods
  findings:
  - statement: Nuclear localization assigned by combined IEA methods; confirmed
      experimentally by OsEME1-GFP imaging.
- id: PMID:12869764
  title: Collection, mapping, and annotation of over 28,000 cDNA clones from japonica
    rice.
  findings:
  - statement: A large-scale rice full-length cDNA collection, sequencing, genome-mapping
      and InterPro-based annotation paper.
  - statement: Contains no gene-specific expression-response experiment for EME1; cannot
      support IEP annotations for response to UV, high light or gamma radiation.
- id: PMID:40333587
  title: ESSENTIAL MEIOTIC ENDONUCLEASE 1 is required for chloroplast development and DNA
    repair in rice.
  findings:
  - statement: OsEME1 (LOC_Os04g55500) is a single-copy nuclear gene; the OsEME1-GFP fusion
      localizes to the nucleus.
  - statement: Recombinant OsEME1 and its C-terminal ERCC4-domain fragment directly bind
      and cleave branched DNA substrates (Y12, pre-X12, X12) in vitro; four conserved
      residues are required for activity.
  - statement: OsEME1 interacts with OsMUS81 (Y2H, luciferase complementation, BiFC); the
      OsEME1 ERCC4 domain and the OsMUS81 HhH motif mediate the interaction.
  - statement: oseme1 loss-of-function mutants are hypersensitive to MMS and Zeocin,
      accumulate gamma-H2AX foci and show S/G2 cell-cycle arrest.
  - statement: oseme1 mutants have a striped-albino-leaf / defective-chloroplast phenotype
      enhanced by high light; the chloroplast defect is downstream of nuclear
      genome-maintenance failure.
  - statement: OsEME1 functions in homologous-recombination repair of DNA damage.
- id: PMID:36495065
  title: MUS81 is required for atypical recombination intermediate resolution but not
    crossover designation in rice.
  findings:
  - statement: rice mus81 mutants have wild-type chiasma numbers and normal HEI10 foci;
      MUS81 does not function in crossover designation in rice.
  - statement: MUS81 resolves atypical meiotic recombination intermediates - mus81 zep1 and
      mus81 fancm double mutants show chromosome fragments and bridges.
- id: PMID:37632767
  title: FANCM interacts with the MHF1-MHF2 complex to limit crossover frequency during
    rice meiosis.
  findings:
  - statement: FANCM-dependent interference-insensitive extra crossovers in rice fancm
      mutants are not marked by HEI10 and require MUS81 resolvase for their resolution.
- id: PMID:23584455
  title: Regulation of Mus81-Eme1 Holliday junction resolvase in response to DNA damage.
  findings:
  - statement: In fission yeast, DNA-damage-induced activation of Mus81-Eme1 requires both
      Cdc2(CDK1)- and Rad3(ATR)-dependent phosphorylation of Eme1.
  - statement: Mus81-Eme1 activation links the DNA-damage response to checkpoint/cell-cycle
      progression.
- id: PMID:20971895
  title: RAD5A, RECQ4A, and MUS81 have specific functions in homologous recombination and
    define different pathways of DNA repair in Arabidopsis thaliana.
  findings:
  - statement: Arabidopsis MUS81 and RECQ4A process recombination-induced aberrant
      intermediates during replication; recq4A mus81 double mutants are lethal.
  - statement: MUS81 is required for efficient synthesis-dependent strand annealing (SDSA),
      an HR-mediated DSB repair pathway.
- id: PMID:24008669
  title: Resolving branched DNA intermediates with structure-specific nucleases during
    replication in eukaryotes.
  findings:
  - statement: MUS81-EME1, GEN1/Yen1 and SLX1-SLX4 resolve branched DNA intermediates
      arising from recombination and from blocked/failing replication forks.
- id: PMID:28049740
  title: Resolvase OsGEN1 Mediates DNA Repair by Homologous Recombination.
  findings:
  - statement: Yen1/GEN1 resolvases work together with MUS81-EME1 and SLX1-SLX4 in
      HR-mediated DNA repair to maintain genome stability.
  - statement: rice osgen1 mutants show reduced chiasma frequency and persistent DSBs in
      microspores, indicating overlapping resolvase activities in plants.
- id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
  title: Deep-research report (falcon / Edison Scientific Literature) - functional
    annotation of rice EME1 (Q0J9J6).
  findings:
  - statement: Synthesizes the rice-specific Du et al. 2025 study (Plant Biotechnology
      Journal, DOI 10.1111/pbi.70101, = PMID:40333587) with Arabidopsis and yeast/human
      MUS81 literature, concluding OsEME1 is best annotated as a nuclear, structure-specific
      endonuclease that cleaves branched/junction DNA substrates (Y12, pre-X12, X12)
      consistent with HR/replication intermediates and interacts with OsMUS81 in the nucleus.
  - statement: Quantifies the rice biochemistry - purified OsEME1 binds the Y12 junction
      substrate with Kd = 16.3 +/- 2.2 microM (microscale thermophoresis); the C-terminal
      ERCC4 fragment carries most nuclease activity; conserved residues L439, P487, E490
      and K494 are required (L439V/E490A/K494E abolish binding/cleavage).
  - statement: Independently corroborates the rice mutant phenotypes - oseme1/k48 is
      hypersensitive to MMS (75 microg/mL) and Zeocin (50 microg/mL), shows S-phase
      accumulation and gamma-H2AX-like foci, and has a striped/albino chloroplast phenotype
      (Fv/Fm 0.53 vs 0.72 WT) enhanced by high light, with 970 (normal-light) and 3548
      (high-light) DEGs.
  - statement: Notes the chloroplast defect is most parsimoniously explained by nuclear
      genome maintenance and/or regulation of nuclear chloroplast-biogenesis genes
      (OsEME1 cleaves GLK1/GLK2-derived structured DNA in vitro); no evidence of stable
      chloroplast localization of the OsEME1 protein.
  - statement: Flags as a caution that in the rice study OsMUS81 bound but did not cleave
      X-structure substrates while OsEME1 produced cleavage products - deviating from the
      canonical cross-species view that MUS81 supplies the catalytic site; to be interpreted
      cautiously and treated as conserved-pathway inference where direct rice data are absent.
core_functions:
- description: >
    OsEME1 is the EME1/MMS4-family subunit of a structure-specific DNA endonuclease that,
    together with MUS81, binds and cleaves branched DNA intermediates - nicked Holliday
    junctions, crossover junctions, 3'-flaps, D-loops and replication-fork structures. In
    rice, recombinant OsEME1 (via its C-terminal ERCC4 domain) directly cleaves
    Holliday-junction-type substrates.
  molecular_function:
    id: GO:0008821
    label: crossover junction DNA endonuclease activity
  directly_involved_in:
  - id: GO:0000724
    label: double-strand break repair via homologous recombination
  locations:
  - id: GO:0005634
    label: nucleus
  in_complex:
    id: GO:0048476
    label: Holliday junction resolvase complex
  supported_by:
  - reference_id: PMID:40333587
    supporting_text: "OsEME1 directly binds to and cleaves Y12, pre-X12 and X12, which are
      typical substrates after HR repair of DNA damage"
  - reference_id: PMID:40333587
    supporting_text: "OsEME1 interacts with OsMUS81 and ... the ERCC4 domain of OsEME1 is
      required for this interaction"
  - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
    supporting_text: "OsEME1 is best annotated as a **nuclear, structure-specific
      endonuclease** that cleaves **branched/junction DNA substrates** consistent with
      HR/replication intermediates, and that interacts with **OsMUS81** in the nucleus."
- description: >
    OsEME1 maintains nuclear genome stability by resolving homologous-recombination
    intermediates and processing stalled/collapsed replication forks during somatic DNA
    repair. Loss of OsEME1 causes hypersensitivity to DNA-damaging agents, gamma-H2AX
    accumulation and cell-cycle arrest, and - as a downstream consequence of genome-
    maintenance failure - defective chloroplast development.
  molecular_function:
    id: GO:0008821
    label: crossover junction DNA endonuclease activity
  directly_involved_in:
  - id: GO:0006302
    label: double-strand break repair
  - id: GO:0031297
    label: replication fork processing
  locations:
  - id: GO:0005634
    label: nucleus
  supported_by:
  - reference_id: PMID:40333587
    supporting_text: "The ... and ... mutants exhibited significantly more γ2HAX foci
      than KY131 with or without Zeocin treatment ... OsEME1 regulates DSB repair and the
      cell cycle"
  - reference_id: PMID:20971895
    supporting_text: "RECQ4A and MUS81 are required for processing recombination-induced
      aberrant intermediates during replication"
  - reference_id: file:ORYSJ/EME1/EME1-deep-research-falcon.md
    supporting_text: "OsEME1 is required to prevent accumulation of DNA damage and
      cell-cycle defects and is necessary for normal chloroplast development and
      photosynthetic performance—likely via a nuclear genome maintenance mechanism
      and/or maintenance of expression of chloroplast-development regulators."
proposed_new_terms: []
suggested_questions:
- question: Does rice OsEME1 have intrinsic catalytic activity, or is its in vitro cleavage
    activity dependent on co-purified or contaminating MUS81 - given that yeast and human
    EME1 are non-catalytic regulatory subunits?
  experts:
  - Guijie Du
- question: What is the relative contribution of OsEME1/OsMUS81 versus OsGEN1 to somatic
    versus meiotic Holliday junction resolution in rice?
  experts:
  - Zhukuan Cheng
- question: Is the striped-leaf/chloroplast phenotype of oseme1 entirely explained by
    nuclear genome-maintenance failure, or does OsEME1 also act on the chloroplast genome?
  experts:
  - Guijie Du
suggested_experiments:
- description: Reconstitute the OsEME1-OsMUS81 heterodimer from separately expressed,
    highly purified subunits and compare nuclease activity on Holliday-junction, 3'-flap
    and replication-fork substrates with each subunit alone, to determine whether OsEME1
    is catalytic or regulatory.
  hypothesis: OsEME1 is the regulatory subunit and MUS81 the catalytic subunit, as in
    yeast and humans; apparent OsEME1-alone activity reflects the heterodimer being the
    functional unit.
  experiment_type: in vitro reconstituted endonuclease assay
- description: Generate oseme1 osgen1 double mutants and quantify meiotic chiasmata,
    chromosome fragmentation, pollen viability and somatic DNA-damage sensitivity.
  hypothesis: OsEME1/OsMUS81 and OsGEN1 act in partially redundant Holliday-junction
    resolution pathways, so the double mutant shows synthetic genome-instability defects.
  experiment_type: genetic epistasis analysis
- description: Test oseme1 mutant sensitivity directly to UV-B/UV-C, gamma irradiation and
    interstrand-crosslinking agents (e.g. mitomycin C) and measure HR-repair efficiency
    with a chromosomal recombination reporter.
  hypothesis: OsEME1 is required for repair of replication-blocking and double-strand
    lesions induced by UV, gamma radiation and crosslinkers, consistent with its branched-
    DNA substrate preference.
  experiment_type: genotoxin-sensitivity and recombination-reporter assay

EME1

Gene Description

Existing Annotations Review

Core Functions

References

Suggested Questions for Experts

Suggested Experiments

Deep Research

Falcon

Research report: Functional annotation of rice EME1 (UniProt Q0J9J6; gene EME1; loci Os04g0648700 / LOC_Os04g55500)

0) Target verification (critical disambiguation)

1) Key concepts and definitions (current understanding)

1.1 Structure-selective endonucleases (SSEs)

1.2 MUS81–EME1 (Mms4) complexes

1.3 “Crossover junction endonuclease” and the reaction catalyzed

2) Rice OsEME1: biochemical function, substrates, and mechanism

2.1 Substrate specificity (direct evidence in rice)

2.2 Domain-to-function mapping and essential residues

2.3 Partnership with MUS81 (direct evidence in rice)

2.4 Note on catalytic contribution within the rice MUS81–EME1 pair

3) Biological roles in rice: DNA repair, cell cycle, and chloroplast development

3.1 DNA damage resistance and DDR activation

3.2 Chloroplast development and photosynthesis phenotypes

4) Subcellular localization (where the gene product acts)

5) Pathway context: homologous recombination, replication stress, and junction processing

5.1 Plant MUS81–EME1 functional archetype (comparative plant evidence)

5.2 Rice MUS81 context supports a conserved partnership model

6) Recent developments (prioritizing 2023–2024)

6.1 2023: MUS81 complexes and DNA–protein crosslink (DPC) repair (mechanistic expansion)

6.2 2024: limited plant MUS81–EME1-specific updates in retrieved corpus

7) Current applications and real-world implementations

8) Expert synthesis and interpretation (authoritative analysis)

9) Key statistics and data summary

10) Cited visual evidence (figures)

11) Reference URLs and publication dates (from retrieved sources)

12) Limitations and confidence notes

Artifacts

Citations

📚 Additional Documentation

Notes

EME1 (OsEME1) — Oryza sativa subsp. japonica — research notes

1. What EME1/MUS81 is — conserved biology

DNA-damage-responsive regulation

2. OsEME1 — the primary rice paper (Du et al. 2025, Plant Biotechnol J)

3. Rice OsMUS81 — meiosis (Mu et al. 2022, New Phytol)

4. Other plant context

5. Assessment of the IEP annotations from PMID:12869764

6. Summary of SPKW-retired annotation assessment

📄 View Raw YAML