JOKA2 (M1BJF6), also called NBR1 homolog, is the potato (Solanum tuberosum) member of the plant NBR1/Joka2 family of selective-autophagy cargo receptors - the functional analogue of mammalian p62/SQSTM1 and NBR1. It has the canonical NBR1 modular architecture: an N-terminal PB1 oligomerization domain, a (degenerate) ZZ-type zinc finger, the NBR1/FW region, a C-terminal UBA ubiquitin-binding domain, and an ATG8-interacting motif (AIM/LIR; 817-WDPI-824). JOKA2 functions as an autophagy cargo adaptor that bridges cargo to the autophagosomal membrane by binding the ATG8-family protein ATG8CL through its AIM; binding is abolished by the W821A/I824A AIM mutation (Joka2-AIM) [PMID:26765567]. Through this ATG8CL-coupled selective-autophagy pathway it contributes to plant immunity: overexpression of wild-type Joka2 (but not Joka2-AIM) restricts lesions caused by the oomycete pathogen Phytophthora infestans, while silencing Joka2 increases susceptibility [PMID:26765567]. During infection, Joka2/ATG8CL-labelled defense-related autophagosomes are diverted to the perihaustorial/extrahaustorial membrane to restrict pathogen growth, and the P. infestans RXLR effector PexRD54 antagonises Joka2 by outcompeting it for ATG8CL binding [PMID:26765567, PMID:29932422]. JOKA2 localizes to cytoplasmic ATG8CL-positive autophagosomes and its cargo is ultimately delivered to the vacuole. Its core molecular function is as a selective-autophagy receptor/adaptor (ATG8-family-protein binding, ubiquitin binding via the UBA domain, autophagy cargo adaptor activity); it is NOT an enzyme or a transmembrane transporter. JOKA2 is a genuine autophagy gene, so its UniProt "Autophagy" keyword reflects real biology - in contrast to many SPKW "autophagy" over-annotations - though the most specific terms (macroautophagy / selective autophagy / autophagy cargo adaptor activity) are preferable to the broad parent terms produced by keyword mapping.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0006914 autophagy | IEA GO_REF:0000043 | MODIFY | Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "Autophagy"; snapshot-only, removed from the current GOA release. JOKA2 is a genuine NBR1/p62-family selective-autophagy cargo receptor, so the autophagy keyword is biologically CORRECT, but "autophagy" is the broad parent of the gene's actual macroautophagy/selective-autophagy role. Reason: Unlike the human "autophagy" SPKW over-annotations on unrelated proteins, this is a legitimate autophagy gene: potato Joka2 is "a selective autophagy cargo receptor of Solanaceous plants that also binds ATG8 via an AIM", it physically associates with ATG8CL (lost in the Joka2-AIM mutant), and overexpression of Joka2 (but not Joka2-AIM) "also activates ATG8CL-mediated selective autophagy" [PMID:26765567]. The cited work explicitly states "Joka2/NBR1 mediated selective autophagy pathway contributes to defense against P. infestans" [PMID:29932422]. Removal therefore lost correct biology. However "autophagy" (GO:0006914) is over-broad; because Joka2 acts in autophagosome-mediated (macro)autophagy, the more specific child term "macroautophagy" (GO:0016236) is preferred, with the selective/cargo-receptor aspect captured by the NEW autophagy cargo adaptor activity MF (GO:0160247) below. MODIFY rather than ACCEPT or REMOVE. Proposed replacements: macroautophagy Supporting Evidence: PMID:26765567 Joka2 was reported as a selective autophagy cargo receptor of Solanaceous plants that also binds ATG8 via an AIM PMID:26765567 This indicates that Joka2 also activates ATG8CL-mediated selective autophagy. file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md JOKA2 is a **selective autophagy cargo receptor** that links cargo to ATG8-positive autophagosomes, with a prominent role in **antimicrobial selective autophagy** during oomycete infection. |
| GO:0015031 protein transport | IEA GO_REF:0000043 | MARK AS OVER ANNOTATED | Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keywords "Protein transport" / "Transport"; snapshot-only, removed from the current GOA release. The keyword reflects the cargo-receptor / autophagosome-delivery role of JOKA2 but maps to a generic vesicular protein-transport process that does not capture the autophagy-specific function. Reason: GOA's removal of this annotation was JUSTIFIED. JOKA2 does deliver cargo (and itself) into ATG8-coated autophagosomes that are then carried to the vacuole - "selective-autophagy employs specialized autophagy cargo receptors that bind ATG8 on autophagosome membranes, and recruit specific cargoes to autophagosomes" [PMID:29932422] - so in a loose sense it participates in protein relocation. However, "protein transport" (GO:0015031) is a broad term most strongly associated with secretory/endomembrane and transmembrane protein trafficking; for an autophagy cargo receptor it is uninformative and potentially misleading, implying a generic transport role the protein does not have. The genuine activity is better captured by the autophagy-specific terms: the molecular function "autophagy cargo adaptor activity" (GO:0160247) and the process "macroautophagy" (GO:0016236), both added/proposed in this review. The keyword-derived parent therefore adds nothing once the specific autophagy annotations are present, and its removal is appropriate. Supporting Evidence: PMID:29932422 selective-autophagy employs specialized autophagy cargo receptors that bind ATG8 on autophagosome membranes, and recruit specific cargoes to autophagosomes file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md JOKA2 is a **selective autophagy cargo receptor** that links cargo to ATG8-positive autophagosomes, with a prominent role in **antimicrobial selective autophagy** during oomycete infection. |
| GO:0031410 cytoplasmic vesicle | IEA GO_REF:0000043 | MODIFY | Summary: SPKW (GO_REF:0000043) annotation derived from the UniProt keyword "Cytoplasmic vesicle"; snapshot-only, removed from the current GOA release. JOKA2 associates with cytoplasmic autophagosomes, so the essence is correct, but "cytoplasmic vesicle" is the broad parent of the specific compartment, the autophagosome. Reason: The annotation is not wrong - JOKA2 localizes to cytoplasmic autophagosomes (a type of cytoplasmic vesicle): "JOKA2 localizes to cytoplasmic puncta consistent with autophagosomes and associates with **ATG8CL**-labeled structures in vivo", and the protein co-localizes with GFP:ATG8CL-labelled autophagosomes [PMID:26765567]. The UniProt subcellular-location annotation lists "Cytoplasmic vesicle, autophagosome", and the cell-biology data place Joka2 specifically on ATG8CL-positive autophagosomes. The broad keyword-derived parent "cytoplasmic vesicle" (GO:0031410) should therefore be replaced by the specific, directly supported child term "autophagosome" (GO:0005776), which is independently annotated in current GOA from the Swiss-Prot subcellular-location vocabulary. MODIFY (generalize -> specialize) rather than MARK_AS_OVER_ANNOTATED. Proposed replacements: autophagosome Supporting Evidence: file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md JOKA2 localizes to cytoplasmic puncta consistent with autophagosomes and associates with **ATG8CL**-labeled structures in vivo PMID:26765567 Joka2:RFP colocalizes with GFP:ATG8CL-labelled autophagosomes |
| GO:0005773 vacuole | IEA GO_REF:0000044 | ACCEPT | Summary: IEA annotation from the UniProtKB/Swiss-Prot subcellular-location vocabulary mapping (SL-0272 Vacuole). Duplicates the EXP vacuole annotation below; the autophagy pathway delivers Joka2-associated cargo to the vacuole. Reason: Consistent with the experimentally supported vacuole annotation (EXP, PMID:29932422) and with autophagy biology: autophagosomes "are then carried to the vacuole for recycling" [PMID:29932422]. The vacuole is the terminal degradative compartment for the selective autophagy pathway in which JOKA2 acts. The IEA duplicate of the EXP annotation is acceptable. Supporting Evidence: PMID:29932422 autophagosomes, which are then carried to the vacuole for recycling |
| GO:0005776 autophagosome | IEA GO_REF:0000044 | ACCEPT | Summary: IEA annotation from the UniProtKB/Swiss-Prot subcellular-location vocabulary mapping (SL-0023 Autophagosome). This is the core, directly supported localization of JOKA2. Reason: Strongly supported by direct cell-biology evidence. JOKA2 localizes to cytoplasmic ATG8CL-positive autophagosomes: "Joka2:RFP colocalizes with GFP:ATG8CL-labelled autophagosomes" [PMID:26765567], and "JOKA2 localizes to cytoplasmic puncta consistent with autophagosomes and associates with ATG8CL-labeled structures in vivo". The autophagosome is the precise compartment where the cargo receptor docks onto ATG8 via its AIM; this annotation is also the proposed replacement for the broad retired "cytoplasmic vesicle" SPKW term. Supporting Evidence: PMID:26765567 Joka2:RFP colocalizes with GFP:ATG8CL-labelled autophagosomes file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md JOKA2 localizes to cytoplasmic puncta consistent with autophagosomes and associates with **ATG8CL**-labeled structures in vivo |
| GO:0008270 zinc ion binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: IEA annotation from InterPro (IPR000433, ZZ-type zinc finger). JOKA2 carries a ZZ-type zinc finger (residues 442-492, with predicted Zn-coordinating residues C447/C450/C471/C474), but the domain is annotated as degenerate. Reason: The ZZ-type zinc finger is a genuine structural feature inferred by InterPro and present in the UniProt feature table (ZN_FING 442-492, "ZZ-type; degenerate"; BINDING residues for Zn(2+)), so zinc binding is plausible. In NBR1-family receptors the ZZ domain participates in cargo recognition rather than catalysis. However, the domain is explicitly degenerate in JOKA2, there is no gene-specific experimental evidence for metal binding, and zinc binding is a structural/auxiliary property rather than the protein's core selective-autophagy-receptor function. Retain as a correct but non-core molecular feature. Supporting Evidence: file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md **ZZ** zinc finger and **FW/NBR1** region (cargo-recognition functions) |
| GO:0005773 vacuole | EXP PMID:29932422 Host autophagy machinery is diverted to the pathogen interfa... | ACCEPT | Summary: Experimental (EXP) vacuole localization from the 2018 study. The vacuole is the terminal compartment of the JOKA2/ATG8CL selective-autophagy pathway. Reason: Supported by the UniProt SUBCELLULAR LOCATION annotation ("Vacuole {ECO:0000269|PubMed:29932422}") and consistent with autophagy biology, in which autophagosomes carrying cargo receptors are delivered to the vacuole: "autophagosomes, which are then carried to the vacuole for recycling" [PMID:29932422]. Accept as a core localization for the pathway endpoint. Supporting Evidence: PMID:29932422 autophagosomes, which are then carried to the vacuole for recycling |
| GO:0005515 protein binding | IPI PMID:26765567 An effector of the Irish potato famine pathogen antagonizes ... | MODIFY | Summary: IPI annotation (with UniProtKB:M1C146, ATG8CL) for the physical Joka2-ATG8CL interaction. "Protein binding" is uninformative; the interaction is the defining ATG8-family-protein / autophagy-cargo-adaptor activity of the receptor. Reason: The interaction partner M1C146 is potato ATG8CL, and the binding occurs through the JOKA2 AIM motif: "potato Joka2, but not the AIM mutant, Joka2AIM, associated with ATG8CL", and "Mutation of the predicted AIM in Joka2 (819-WDPI-822) to ADPA resulted in loss of ATG8CL binding" [PMID:26765567]. Per curation guidelines the vague "protein binding" (GO:0005515) should be replaced by an informative molecular function. The most accurate term is "autophagy cargo adaptor activity" (GO:0160247) - the binding activity that brings cargo to the phagophore via ATG8 - capturing both the ATG8-family-protein binding and the receptor/adaptor role. This is also added as a NEW annotation below; MODIFY here to retain the IPI evidence with M1C146. Proposed replacements: autophagy cargo adaptor activity Supporting Evidence: PMID:26765567 potato Joka2, but not the AIM mutant, Joka2AIM, associated with ATG8CL PMID:26765567 Mutation of the predicted AIM in Joka2 (819-WDPI-822) to ADPA resulted in loss of ATG8CL binding |
| GO:0050832 defense response to fungus | IDA PMID:29932422 Host autophagy machinery is diverted to the pathogen interfa... | MODIFY | Summary: IDA annotation for JOKA2's role in defense against the late blight pathogen. The biological conclusion (positive role in pathogen defense) is well supported, but the pathogen is Phytophthora infestans, an OOMYCETE, not a fungus, so the term is taxonomically incorrect. Reason: The defense role is strongly supported: "Overexpression of Joka2, but not Joka2AIM, significantly restricted the size of the disease lesions caused by P. infestans" and "virus-induced gene silencing of Joka2 resulted in increased disease lesions", leading the authors to conclude "Joka2-mediated selective autophagy contributes to defense against this pathogen" [PMID:26765567]; the 2018 study reiterates that "Joka2/NBR1 mediated selective autophagy pathway contributes to defense against P. infestans" [PMID:29932422]. However, Phytophthora infestans is an oomycete (Stramenopiles), not a true fungus, so "defense response to fungus" (GO:0050832) is the wrong taxonomic branch. It should be MODIFIED to "defense response to oomycetes" (GO:0002229), the accurate term, retaining the IDA evidence. Proposed replacements: defense response to oomycetes Supporting Evidence: PMID:26765567 Overexpression of Joka2, but not Joka2AIM, significantly restricted the size of the disease lesions caused by P. infestans PMID:29932422 we recently showed that Joka2/NBR1 mediated selective autophagy pathway contributes to defense against P. infestans |
| GO:0160247 autophagy cargo adaptor activity | IPI PMID:26765567 An effector of the Irish potato famine pathogen antagonizes ... | NEW | Summary: JOKA2 is a selective-autophagy cargo receptor that bridges cargo to the autophagosomal membrane by binding the ATG8-family protein ATG8CL through its AIM motif. This precise molecular function is not represented in current GOA (only the vague "protein binding"). Reason: The current/seeded MF annotation is only the uninformative "protein binding" (GO:0005515). The defining activity of JOKA2 is autophagy cargo adaptor activity (GO:0160247) - "the binding activity of a molecule that brings together a cargo, targeted for degradation via autophagy, to a phagophore". JOKA2 "binds ATG8 via an AIM" and is "a selective autophagy cargo receptor of Solanaceous plants" [PMID:26765567]; the AIM-dependent ATG8CL interaction (lost in Joka2-AIM) is required for its function as a host autophagy cargo receptor (UniProt DOMAIN). IPI is justified by the in planta co-immunoprecipitation of Joka2 with ATG8CL (UniProtKB:M1C146) and the AIM-mutant loss of binding. Supporting Evidence: PMID:26765567 Joka2 was reported as a selective autophagy cargo receptor of Solanaceous plants that also binds ATG8 via an AIM PMID:26765567 potato Joka2, but not the AIM mutant, Joka2AIM, associated with ATG8CL file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md Joka2 binds **ATG8CL** through an **AIM**, and this binding is required for functional immune output |
| GO:0043130 ubiquitin binding | ISS GO_REF:0000043 | NEW | Summary: JOKA2 has a C-terminal UBA domain (residues 811-860) characteristic of NBR1/p62-family receptors, which bind ubiquitin to recognize ubiquitinated cargo. Inferred from sequence/ domain similarity; not yet directly demonstrated for the potato protein. Reason: NBR1/p62-family selective-autophagy receptors couple ubiquitinated cargo to ATG8 via a C-terminal UBA ubiquitin-binding domain. JOKA2 has this UBA domain (UniProt DOMAIN 811-860; Pfam PF24932 UBA_NBR1_C, PROSITE UBA), and the deep-research synthesis notes that "the receptor's UBA domains strongly imply ubiquitin-binding capability, consistent with plant NBR1 family behavior". Ubiquitin binding (GO:0043130) is therefore a strongly supported family-level molecular function. Marked ISS because, although the domain is present, direct ubiquitin-binding has not been shown for potato JOKA2 specifically (it is established for the Arabidopsis/general plant NBR1 family). Provided as a NEW, conservative MF. Supporting Evidence: file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md the receptor’s **UBA domains** strongly imply ubiquitin-binding capability, consistent with plant NBR1 family behavior file:SOLTU/JOKA2/JOKA2-deep-research-falcon.md **UBA** domain(s) (ubiquitin binding) |
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Download this section (compressed HTML)Q: Does potato JOKA2 mediate aggrephagy (GO:0035973, selective autophagy of protein aggregates) via PB1-driven oligomerization and cargo condensation, as inferred from the plant NBR1/Joka2 family? (Both ubiquitin binding, GO:0043130, and macroautophagy/cargo-adaptor activity are already captured above; aggrephagy is the family-level process still untested in potato.)
Suggested experts: Yan Zhang
Q: What are the endogenous defense-related cargoes that potato JOKA2 delivers to autophagosomes during P. infestans infection, and are they ubiquitinated?
Suggested experts: Tolga O. Bozkurt
Q: Does potato JOKA2 recognize non-ubiquitinated cargo via its ZZ+FW region and undergo liquid-liquid phase separation, as recently shown for Arabidopsis NBR1?
Suggested experts: Yasin F. Dagdas
Q: Is JOKA2's UBA domain functional for ubiquitin binding in potato, and how does ubiquitin-dependent versus ATG8-dependent recruitment partition its activity?
Suggested experts: Sophien Kamoun
Experiment: Identify endogenous JOKA2 cargo by affinity-purification mass spectrometry of Joka2 versus Joka2-AIM during P. infestans infection, and test ubiquitination of candidate cargoes.
Hypothesis: JOKA2 selectively captures ubiquitinated defense-related or pathogen-derived proteins for autophagic delivery to the vacuole.
Type: affinity-purification proteomics
Experiment: Test recombinant JOKA2 UBA domain for binding to mono- and poly-ubiquitin chains in vitro (ITC/pulldown), and assess cargo recruitment in planta with a UBA-domain mutant.
Hypothesis: The JOKA2 UBA domain binds ubiquitin and is required for recognition of ubiquitinated cargo, as in other NBR1/p62-family receptors.
Type: in vitro ubiquitin-binding and structure-function assay
Experiment: Quantify P. infestans colonization and autophagic flux in joka2 loss-of-function potato lines complemented with wild-type, AIM-mutant, UBA-mutant or PB1-mutant JOKA2.
Hypothesis: ATG8 binding (AIM), ubiquitin binding (UBA) and oligomerization (PB1) each contribute to JOKA2-mediated selective autophagy and immunity against the oomycete.
Type: genetic complementation and disease-resistance assay
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