Cytochrome P450 71A13, an N-terminally membrane-anchored heme-thiolate monooxygenase that carries out the second step of camalexin biosynthesis in Arabidopsis. Using NADPH-P450 reductase, it converts indole-3-acetaldoxime (IAOx, made by CYP79B2/B3) to indole-3-acetonitrile (IAN) and, oxidatively, to an indole cyanohydrin/dehydro-IAN electrophile that is captured by glutathione or cysteine en route to camalexin, which CYP71B15 (PAD3) completes. It is the predominant isoform for this step in leaves; its tandem paralog CYP71A12 is partially redundant and mainly channels IAN towards indole-3-carboxylic acid derivatives. CYP71A13 forms a membrane-associated metabolon with CYP79B2, CYP71B15, ATR1 and GSTU4. The gene is strongly induced by pathogens, elicitors and UV-C, and mutants are camalexin-deficient and susceptible to necrotrophic fungi such as Alternaria brassicicola.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004497 monooxygenase activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro P450 mapping to monooxygenase activity. Correct: CYP71A13 is an NADPH/P450-reductase-dependent heme-thiolate monooxygenase whose IAOx turnover is oxidative. Reason: Klein et al. 2013 showed IAOx turnover by CYP71A13 is strictly NADPH-dependent and proceeds via oxidation (proposed alpha-hydroxy-IAN/dehydro-IAN), so the generic monooxygenase term is supported beyond the domain mapping. The specific activity is better captured by GO:0047720, but this broader IEA is accurate. 18O2 labeling shows oxygen incorporation from O2 into the IAL product, so a genuine monooxygenation occurs. (The independent review kept this row as non-core because the family-level term is less informative than GO:0047720; retained as ACCEPT here since it is accurate and supported beyond the domain mapping.) Supporting Evidence: PMID:24151049 With 97% 18O2, both CYP71A13 and CYP71A12 catalyze incorporation of the isotopic label into IAL PMID:24151049 the formation of both IAN and IAL is strictly dependent on the presence of both CYP71A13 and the cofactor NADPH PMID:24151049 These data establish that CYP71A13 catalyzes the conversion of IAOx to an electrophile capable of accepting the thiol group of L-cysteine or glutathione |
| GO:0005506 iron ion binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: Iron ion binding mapped from P450 InterPro signatures; reflects the heme iron. Reason: Iron is bound only as heme iron (heme-thiolate Cys439); this is a cofactor-level descriptor rather than a function, redundant with heme binding. Not wrong, retained as non-core. Supporting Evidence: file:ARATH/CYP71A13/CYP71A13-uniprot.txt SIMILARITY: Belongs to the cytochrome P450 family. |
| GO:0005515 protein binding | IPI PMID:19452453 Proteomic profiling of tandem affinity purified 14-3-3 prote... | REMOVE | Summary: CYP71A13 recovered as a putative client in TAP-tag purification of 14-3-3 omega (GRF2, At1g78300) complexes. Reason: Generic protein binding is uninformative. This is a high-throughput 14-3-3 client screen; no functional consequence for CYP71A13 was tested and no more specific MF is supported. Removal does not imply the interaction is false. |
| GO:0005515 protein binding | IPI PMID:31511315 The Formation of a Camalexin Biosynthetic Metabolon. | REMOVE | Summary: CYP71A13 interacts with CYP71B15 (PAD3) by co-IP and FRET-FLIM in the camalexin metabolon. Reason: Generic protein binding is uninformative. The interaction is real and is captured functionally by the catalytic complex (GO:1902494) and camalexin biosynthetic process annotations; no more specific MF is warranted from the interaction alone. Removal does not imply the interaction is false. Supporting Evidence: PMID:31511315 we used two independent untargeted coimmunoprecipitation (co-IP) approaches with the biosynthetic enzymes CYP71B15 and CYP71A13 as baits and determined that the camalexin biosynthetic P450 enzymes copurified with these enzymes |
| GO:0005515 protein binding | IPI PMID:31511315 The Formation of a Camalexin Biosynthetic Metabolon. | REMOVE | Summary: CYP71A13 interacts with GSTU4, recruited to the camalexin metabolon. Reason: Generic protein binding is uninformative. The interaction is real and is captured functionally by the catalytic complex (GO:1902494) and camalexin biosynthetic process annotations; no more specific MF is warranted from the interaction alone. Removal does not imply the interaction is false. Supporting Evidence: PMID:31511315 GSTU4, which is coexpressed with Trp- and camalexin-specific enzymes, is physically recruited to the complex |
| GO:0005739 mitochondrion | ISM GO_REF:0000122 | REMOVE | Summary: AtSubP sequence-based prediction of mitochondrial localization. Reason: CYP71A13 has an N-terminal hydrophobic signal anchor (TM 2-20) typical of ER-anchored class II plant P450s that are isolated in microsomes and partner with the ER P450 reductase ATR1; there is no experimental support for mitochondrial targeting. The prediction is contradicted by the protein architecture. Supporting Evidence: file:ARATH/CYP71A13/CYP71A13-uniprot.txt SUBCELLULAR LOCATION: Membrane {ECO:0000305}; Single-pass membrane PMID:24151049 these membrane-associated enzymes were isolated in the microsomal fraction |
| GO:0005788 endoplasmic reticulum lumen | IDA PMID:33831160 The Formation of a Camalexin Biosynthetic Metabolon. | MODIFY | Summary: TAIR IDA (2022-02-03) for ER lumen. The cited PMID:33831160 is a deleted duplicate of PMID:31511315 (Mucha et al. 2019), whose evidence is confocal colocalization of CYP71A13 fusions with the ER lumenal marker RFP-HDEL. That shows ER localization but cannot distinguish the lumen from the membrane. Reason: The ER localization is experimentally sound and is kept. The lumen term is not: CYP71A13 is a signal-anchored class II P450 with a single N-terminal transmembrane helix, and the same paper states that such P450s face the cytosol. Mucha et al. 2019 (PMID:31511315; full text read on PMC, PMC6881122, not cacheable) states "Typically, eukaryotic P450 enzymes are anchored to the membrane of the endoplasmic reticulum (ER) with their catalytic centers facing the cytosolic side" and reports that CYP71A12, CYP71A13 and CYP71B15 "were localized to the ER and showed colocalization with the ER lumenal marker RFP-HDEL" (Supplemental Figure 4). Colocalization with a lumenal marker at confocal resolution identifies the organelle, not the side of the membrane, so endoplasmic reticulum membrane is the correct term. The Mucha full text could not be cached (PMC returns abstract only), so its sentences are quoted here rather than in supported_by. Proposed replacements: endoplasmic reticulum membrane Supporting Evidence: file:ARATH/CYP71A13/CYP71A13-uniprot.txt SUBCELLULAR LOCATION: Membrane {ECO:0000305}; Single-pass membrane PMID:24151049 these membrane-associated enzymes were isolated in the microsomal fraction file:ARATH/CYP71A13/CYP71A13-deep-research-falcon.md Fluorescent CYP71A13 fusions showed ER localization when expressed in file:ARATH/CYP71A13/CYP71A13-deep-research-openscientist.md where it is reduced by the ER-resident NADPH–cytochrome P450 reductase |
| GO:0006952 defense response | NAS PMID:31511315 The Formation of a Camalexin Biosynthetic Metabolon. | KEEP AS NON CORE | Summary: ComplexPortal NAS for defense response, from the camalexin metabolon paper. Reason: Correct but very general; CYP71A13 contributes to defense by producing the phytoalexin camalexin. More specific defense-response-to-fungus annotations exist. Retained as non-core. Supporting Evidence: PMID:17573535 Plants carrying cyp71A13 mutations produce greatly reduced amounts of camalexin after infection by Pseudomonas syringae or A. brassicicola and are susceptible to A. brassicicola PMID:31511315 we used two independent untargeted coimmunoprecipitation (co-IP) approaches with the biosynthetic enzymes CYP71B15 and CYP71A13 as baits and determined that the camalexin biosynthetic P450 enzymes copurified with these enzymes |
| GO:0009617 response to bacterium | IEP PMID:17573535 Arabidopsis cytochrome P450 monooxygenase 71A13 catalyzes th... | KEEP AS NON CORE | Summary: Expression of CYP71A13 is induced by bacterial infection (P. syringae). Reason: Transcriptional induction by pathogens is well documented (UniProt INDUCTION), and IEP is the appropriate evidence for a response term. It describes regulation of the gene rather than its function; non-core. Supporting Evidence: file:ARATH/CYP71A13/CYP71A13-uniprot.txt INDUCTION: By Pseudomonas syringae pv. tomato DC3000, Botrytis cinerea PMID:17573535 Plants carrying cyp71A13 mutations produce greatly reduced amounts of camalexin after infection by Pseudomonas syringae or A. brassicicola and are susceptible to A. brassicicola |
| GO:0009617 response to bacterium | IMP PMID:23073694 Metabolic and transcriptomic changes induced in Arabidopsis ... | KEEP AS NON CORE | Summary: cyp71A13 mutant tested among camalexin/glucosinolate mutants in rhizobacterium (P. fluorescens SS101)-induced resistance to Pst. Reason: Mutant phenotype reflects loss of camalexin, which is required for SS101-induced resistance. CYP71A13 acts upstream via phytoalexin synthesis; a non-core, indirect process. Supporting Evidence: PMID:23073694 showed that camalexin and glucosinolates are indeed required for the induction of Pst resistance by Pf .SS101 |
| GO:0009682 induced systemic resistance | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: PAINT IBA for induced systemic resistance at node PTN005384924, grounded in CYP71A12 and CYP71A13 themselves. Reason: The node is the recent CYP71A12/CYP71A13 clade and the IBD reflects the experimental IMP for both paralogs (target appearing in its own WITH/FROM is expected), but those rows are themselves marked over-annotated. GO:0009682 is defined as a response to non-pathogenic bacteria conferring systemic resistance "that does not depend upon salicylic acid signaling", whereas the only experimental source (van de Mortel et al. 2012) states that the Pf.SS101-induced resistance to Pst is salicylic acid-dependent and not jasmonic acid/ethylene-dependent. The term therefore misdescribes the observed response. The requirement of camalexin for this resistance is captured by defense response to bacterium. Propagation Review Root cause: SOURCE BAD Sources checked: AGI_LocusCode:AT2G30750 · CYP71A12 SOURCE BAD IMP from van de Mortel et al. 2012; the observed resistance is salicylic acid-dependent, outside the GO:0009682 definition. AGI_LocusCode:AT2G30770 · CYP71A13 SOURCE BAD Same paper and same definitional mismatch as CYP71A12. PANTHER:PTN005384924 · PTN005384924 SOURCE BAD The node's only experimental grounding is the two IMP rows above. Supporting Evidence: PMID:23073694 showed that camalexin and glucosinolates are indeed required for the induction of Pst resistance by Pf .SS101 PMID:23073694 the Pf .SS101 -induced resistance response to Pst is dependent on salicylic acid signaling and not on jasmonic acid and ethylene signaling |
| GO:0009682 induced systemic resistance | IMP PMID:23073694 Metabolic and transcriptomic changes induced in Arabidopsis ... | MARK AS OVER ANNOTATED | Summary: cyp71A13 mutant impaired in P. fluorescens SS101-induced resistance against Pst. Reason: The mutant phenotype (via camalexin biosynthesis) is real, but the term does not fit it. GO:0009682 is defined as a response to non-pathogenic bacteria conferring systemic resistance "that does not depend upon salicylic acid signaling", whereas the only experimental source (van de Mortel et al. 2012) states that the Pf.SS101-induced resistance to Pst is salicylic acid-dependent and not jasmonic acid/ethylene-dependent. The term therefore misdescribes the observed response. The same treatment is applied to the identical rows on CYP71A12 and MYB51. The requirement of CYP71A13 for resistance to Pst is captured by defense response to bacterium. Supporting Evidence: PMID:23073694 showed that camalexin and glucosinolates are indeed required for the induction of Pst resistance by Pf .SS101 PMID:23073694 the Pf .SS101 -induced resistance response to Pst is dependent on salicylic acid signaling and not on jasmonic acid and ethylene signaling |
| GO:0010120 camalexin biosynthetic process | IMP PMID:17573535 Arabidopsis cytochrome P450 monooxygenase 71A13 catalyzes th... | ACCEPT | Summary: cyp71A13 mutants are strongly reduced in camalexin and are rescued by IAN feeding. Reason: Core process. CYP71A13 catalyzes the IAOx conversion step in camalexin biosynthesis; the single mutant retains ~2% camalexin and the cyp71a12 cyp71a13 double knockout is essentially camalexin-free. The enzyme performs a step of the pathway, so involved_in is warranted. Supporting Evidence: PMID:17573535 Plants carrying cyp71A13 mutations produce greatly reduced amounts of camalexin after infection by Pseudomonas syringae or A. brassicicola and are susceptible to A. brassicicola PMID:17573535 Exogenously supplied IAN restored camalexin production in cyp71A13 mutant plants PMID:25953104 In cyp71a12 cyp71a13 double mutants, only traces of camalexin are synthesized, showing that dehydration of IAOx by CYP71A12 and CYP71A13 is essential for IAN synthesis as a camalexin precursor. |
| GO:0010120 camalexin biosynthetic process | NAS PMID:31511315 The Formation of a Camalexin Biosynthetic Metabolon. | ACCEPT | Summary: ComplexPortal NAS for camalexin biosynthesis from the metabolon paper. Reason: Consistent with genetic and biochemical evidence; CYP71A13 is the dominant leaf enzyme for the IAOx step and forms a metabolon with CYP79B2 and CYP71B15. Supporting Evidence: PMID:31511315 we used two independent untargeted coimmunoprecipitation (co-IP) approaches with the biosynthetic enzymes CYP71B15 and CYP71A13 as baits and determined that the camalexin biosynthetic P450 enzymes copurified with these enzymes PMID:31511315 indole-3-cyanohydrin, which is synthesized by CYP71A12 and especially CYP71A13 PMID:24151049 These data establish that CYP71A13 catalyzes the conversion of IAOx to an electrophile capable of accepting the thiol group of L-cysteine or glutathione |
| GO:0016020 membrane | IEA GO_REF:0000044 | ACCEPT | Summary: Membrane from UniProt subcellular location (single-pass membrane protein). Reason: Correct but general; CYP71A13 is N-terminally anchored and microsomal. The specific compartment, endoplasmic reticulum membrane (GO:0005789), is captured by the MODIFY of the ER lumen IDA from Mucha et al. 2019, which shows ER localization by confocal colocalization with an ER marker. Supporting Evidence: file:ARATH/CYP71A13/CYP71A13-uniprot.txt SUBCELLULAR LOCATION: Membrane {ECO:0000305}; Single-pass membrane PMID:24151049 these membrane-associated enzymes were isolated in the microsomal fraction |
| GO:0016020 membrane | NAS PMID:31511315 The Formation of a Camalexin Biosynthetic Metabolon. | ACCEPT | Summary: ComplexPortal NAS: membrane. Reason: Correct; the camalexin metabolon is a membrane-associated P450 complex. Supporting Evidence: PMID:31511315 we used two independent untargeted coimmunoprecipitation (co-IP) approaches with the biosynthetic enzymes CYP71B15 and CYP71A13 as baits and determined that the camalexin biosynthetic P450 enzymes copurified with these enzymes |
| GO:0016491 oxidoreductase activity | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT IBA for oxidoreductase activity at a deep CYP71 node. Reason: Correct, if general, for a P450 monooxygenase whose IAOx turnover is NADPH-dependent and oxidative. Supporting Evidence: PMID:24151049 the formation of both IAN and IAL is strictly dependent on the presence of both CYP71A13 and the cofactor NADPH |
| GO:0016705 oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro mapping to paired-donor oxygen-incorporating oxidoreductase activity (P450 class). Reason: Appropriate class for an NADPH-P450 reductase-dependent P450; the oxidative IAOx turnover supports it. Supporting Evidence: PMID:24151049 the formation of both IAN and IAL is strictly dependent on the presence of both CYP71A13 and the cofactor NADPH |
| GO:0020037 heme binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: Heme binding from P450 InterPro signatures. Reason: Correct cofactor binding (heme-thiolate Cys439) shared by all P450s; descriptive of the fold rather than the specific function. Supporting Evidence: file:ARATH/CYP71A13/CYP71A13-uniprot.txt SIMILARITY: Belongs to the cytochrome P450 family. |
| GO:0042742 defense response to bacterium | IMP PMID:17573535 Arabidopsis cytochrome P450 monooxygenase 71A13 catalyzes th... | KEEP AS NON CORE | Summary: cyp71A13 mutant phenotype in defense against bacteria. Reason: cyp71A13 mutants make little camalexin after P. syringae infection; the defense contribution is indirect (via the phytoalexin) and the clearest pathogen phenotype is against fungi. Rajniak et al. 2015 independently reported that cyp71A13 mutants are more susceptible to P. syringae. Retained as non-core, acts upstream. Supporting Evidence: PMID:17573535 Plants carrying cyp71A13 mutations produce greatly reduced amounts of camalexin after infection by Pseudomonas syringae or A. brassicicola and are susceptible to A. brassicicola PMID:26352477 Notably, the adult leaves and seedlings of the camalexin pathway mutants cyp71A13 and pad3 are also more susceptible to Pst infection compared to WT |
| GO:0047720 indoleacetaldoxime dehydratase activity | IDA PMID:17573535 Arabidopsis cytochrome P450 monooxygenase 71A13 catalyzes th... | ACCEPT | Summary: Recombinant CYP71A13 converts IAOx to IAN (EC 4.8.1.3). Reason: Core MF, directly demonstrated by Nafisi et al. 2007 and reproduced by Klein et al. 2013 and Muller et al. 2015. Note the reaction is NADPH-dependent and mechanistically oxidative (cyanohydrin/dehydro-IAN intermediates that are captured by Cys/GSH), so this dehydratase term describes the net IAN-forming outcome; no GO term yet describes the oxidative coupling-competent step. Supporting Evidence: PMID:17573535 CYP71A13 expressed in Escherichia coli converted IAOx to indole-3-acetonitrile (IAN) PMID:24151049 the formation of both IAN and IAL is strictly dependent on the presence of both CYP71A13 and the cofactor NADPH PMID:24151049 These data establish that CYP71A13 catalyzes the conversion of IAOx to an electrophile capable of accepting the thiol group of L-cysteine or glutathione |
| GO:0047720 indoleacetaldoxime dehydratase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Rhea/EC-based IEA (RHEA:23156, EC 4.8.1.3) for indoleacetaldoxime dehydratase activity. Reason: Consistent with the experimental IDA for the same activity. Supporting Evidence: PMID:17573535 CYP71A13 expressed in Escherichia coli converted IAOx to indole-3-acetonitrile (IAN) |
| GO:0050832 defense response to fungus | IMP PMID:17573535 Arabidopsis cytochrome P450 monooxygenase 71A13 catalyzes th... | ACCEPT | Summary: cyp71A13 mutants are susceptible to Alternaria brassicicola. Reason: Strong and reproducible: susceptibility to A. brassicicola, loss of UV-C-induced resistance to B. cinerea, and immunity defects with P. cucumerina/C. tropicale. The enzyme acts upstream of fungal defense by producing camalexin; this is the main physiological role of the gene. Supporting Evidence: PMID:17573535 Plants carrying cyp71A13 mutations produce greatly reduced amounts of camalexin after infection by Pseudomonas syringae or A. brassicicola and are susceptible to A. brassicicola PMID:19154205 UV-C-induced resistance was not seen in the camalexin-deficient mutants cyp79B2/B3, cyp71A13, pad3 or pad2 PMID:31411742 revealed cyp71A12-, cyp71A13- and cyp82C2-associated defects associated with Arabidopsis immunity |
| GO:1902494 catalytic complex | NAS PMID:31511315 The Formation of a Camalexin Biosynthetic Metabolon. | KEEP AS NON CORE | Summary: ComplexPortal NAS: part of the camalexin biosynthetic metabolon (catalytic complex). Reason: Mucha et al. 2019 showed by co-IP and FRET-FLIM that CYP71A13 associates with CYP79B2, CYP71B15, ATR1 and GSTU4, and allosterically increases CYP79B2 substrate affinity. The association is real and catalytic complex is the only available CC term, but the assembly is a dynamic, pathogen-induced metabolon rather than a stable complex of fixed composition, and CYP71A13 is catalytically active on its own with P450 reductase (Klein et al. 2013). Kept as non-core, consistent with the CYP71A12, CYP71B15 and CYP79B2 reviews. Supporting Evidence: PMID:31511315 we used two independent untargeted coimmunoprecipitation (co-IP) approaches with the biosynthetic enzymes CYP71B15 and CYP71A13 as baits and determined that the camalexin biosynthetic P450 enzymes copurified with these enzymes PMID:31511315 GSTU4, which is coexpressed with Trp- and camalexin-specific enzymes, is physically recruited to the complex PMID:31511315 We detected increased substrate affinity of CYP79B2 in the presence of CYP71A13 PMID:24151049 the formation of both IAN and IAL is strictly dependent on the presence of both CYP71A13 and the cofactor NADPH |
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Download this section (compressed HTML)Q: Should GO add a molecular function term for the NADPH-dependent oxidative conversion of indole-3-acetaldoxime to the thiol-reactive cyanohydrin/dehydro-IAN intermediate, distinct from EC 4.8.1.3 dehydratase activity?
Q: Should TAIR revise the ER lumen (GO:0005788) IDA rows from Mucha et al. 2019 (cited under the deleted duplicate PMID:33831160) on CYP71A12, CYP71A13 and CYP71B15 to endoplasmic reticulum membrane, given that the evidence is colocalization with a lumenal marker and the paper itself describes these P450s as membrane-anchored and cytosol-facing?
Q: How much does CYP71A12 compensate for CYP71A13 in camalexin synthesis in different tissues, given that CYP71A12 favours the indole-3-carbaldehyde/indole-3-carboxylic acid branch?
Q: Is conjugation of the CYP71A13 product to glutathione enzyme-catalyzed in planta (GSTF6 or another GST), given that metabolon-recruited GSTU4 antagonizes camalexin accumulation?
Q: Does the vasculature-restricted induction of CYP71A13 versus epidermal induction of CYP71A12 at fungal infection sites (Tang et al. 2023) reflect distinct metabolic outputs (camalexin vs ICOOH derivatives) per cell type?
Experiment: Co-localize CYP71A13-FP with ER markers and test protease protection / glycosylation-site topology to resolve the ER lumen annotation.
Hypothesis: CYP71A13 is ER-membrane anchored with a cytosol-facing catalytic domain.
Type: subcellular localization and topology
Experiment: Complement cyp71a12 cyp71a13 double mutants with CYP71A13 driven by vasculature- versus epidermis-specific promoters and score camalexin and C. higginsianum colonization.
Hypothesis: Cell-type-specific CYP71A13 expression in vasculature is required for full camalexin-mediated resistance to Colletotrichum.
Type: cell-type-specific complementation
Experiment: Kinetic comparison of purified CYP71A13 and CYP71A12 with IAOx and IAN, with and without glutathione transferases, to define how product partitioning between Cys/GS-IAN and indole-3-carbaldehyde is controlled.
Hypothesis: Product partitioning between Cys/GS-IAN and indole-3-carbaldehyde differs intrinsically between CYP71A13 and CYP71A12.
Type: in vitro enzyme kinetics
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