ClpX (CG4538) encodes the AAA+ ATPase subunit of the Drosophila mitochondrial ClpXP protease. Hexameric ClpX recognizes specific matrix proteins, unfolds them with energy from ATP hydrolysis and translocates them into the proteolytic chamber of the ClpP tetradecamer. In fly cells ClpX binds both ClpP and the mitochondrial RNA-binding protein DmLRPPRC1, and loss of ClpX or ClpP stabilizes DmLRPPRC1, linking ClpXP-mediated proteolysis to control of mitochondrial mRNA abundance and translation and to mitochondrial protein quality control.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005524 ATP binding | IBA GO_REF:0000033 | ACCEPT | Summary: ATP binding inherited from the ClpX ancestral node. Reason: ClpX is an AAA+ ATPase; nucleotide binding is required for its motor function. Supporting Evidence: PMID:28814717 ClpXP is composed of a proteolytic subunit, ClpP, and a chaperone-like subunit, ClpX, which carries an ATPase associated with diverse cellular activities (AAA+) domain6, 7. |
| GO:0005524 ATP binding | IEA GO_REF:0000002 | ACCEPT | Summary: ATP binding from AAA+ and ClpX InterPro signatures. Reason: Same as above. Supporting Evidence: PMID:28814717 ClpXP is composed of a proteolytic subunit, ClpP, and a chaperone-like subunit, ClpX, which carries an ATPase associated with diverse cellular activities (AAA+) domain6, 7. |
| GO:0005759 mitochondrial matrix | IBA GO_REF:0000033 | ACCEPT | Summary: Mitochondrial matrix inherited from the mitochondrial ClpX ancestral node. Reason: Fly ClpX belongs to the mitochondrial ClpX clade and partners with mitochondrial ClpP. Supporting Evidence: PMID:28814717 ClpXP and Lon localize to the matrix space whereas the m-AAA protease is anchored in the membrane with its catalytic site exposed to the matrix space. |
| GO:0005759 mitochondrial matrix | IC PMID:28814717 Drosophila protease ClpXP specifically degrades DmLRPPRC1 co... | ACCEPT | Summary: Mitochondrial matrix inferred from mitochondrial Clp complex membership. Reason: ClpXP is a matrix protease. Supporting Evidence: PMID:28814717 ClpXP and Lon localize to the matrix space whereas the m-AAA protease is anchored in the membrane with its catalytic site exposed to the matrix space. |
| GO:0006457 protein folding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: Protein folding from the ClpX InterPro signature. Reason: ClpX primarily unfolds substrates for ClpP degradation; mammalian and yeast mitochondrial ClpX also remodel some client proteins independently of ClpP, but this has not been tested in flies. Keep as a non-core, family-level inference. Supporting Evidence: PMID:28814717 ClpXP is composed of a proteolytic subunit, ClpP, and a chaperone-like subunit, ClpX, which carries an ATPase associated with diverse cellular activities (AAA+) domain6, 7. PMID:28814717 Moreover, we showed that ClpX, which recognize the protein substrates of ClpXP, physically interacts with DmLRPPRC1 in the cells (Fig. |
| GO:0009841 mitochondrial endopeptidase Clp complex | IDA PMID:28814717 Drosophila protease ClpXP specifically degrades DmLRPPRC1 co... | ACCEPT | Summary: FLAG-tagged ClpX co-immunoprecipitates ClpP and the substrate DmLRPPRC1. Reason: Core complex membership. Supporting Evidence: PMID:28814717 FLAG-tagged ClpX is co-immunoprecipitated with ClpP and DmLRPPRC1 but not DmSLIRP1. |
| GO:0016887 ATP hydrolysis activity | IBA GO_REF:0000033 | ACCEPT | Summary: ATP hydrolysis activity inherited from the ClpX ancestral node. Reason: ClpX is the ATP-hydrolysing AAA+ motor of ClpXP. Supporting Evidence: PMID:28814717 ClpXP is composed of a proteolytic subunit, ClpP, and a chaperone-like subunit, ClpX, which carries an ATPase associated with diverse cellular activities (AAA+) domain6, 7. |
| GO:0016887 ATP hydrolysis activity | IEA GO_REF:0000002 | ACCEPT | Summary: ATP hydrolysis activity from the AAA+ ATPase InterPro signature. Reason: Same as above. Supporting Evidence: PMID:28814717 ClpXP is composed of a proteolytic subunit, ClpP, and a chaperone-like subunit, ClpX, which carries an ATPase associated with diverse cellular activities (AAA+) domain6, 7. |
| GO:0030163 protein catabolic process | IBA GO_REF:0000033 | MODIFY | Summary: Protein catabolic process inherited from the ClpX ancestral node. Reason: ClpX delivers substrates for ClpP-mediated degradation; consistent with fly knockdown data. The fly ClpXP acts in the mitochondrial matrix, so the more specific mitochondrial protein catabolic process applies. Proposed replacements: mitochondrial protein catabolic process Supporting Evidence: PMID:28814717 Here, we showed that knockdown of ClpX or ClpP results in an increase of DmLRPPRC1 protein without an increase in the corresponding mRNA (Figs 2 and S4). |
| GO:0030163 protein catabolic process | IMP PMID:28814717 Drosophila protease ClpXP specifically degrades DmLRPPRC1 co... | MODIFY | Summary: ClpX knockdown stabilizes DmLRPPRC1 protein without changing its mRNA. Reason: Direct fly evidence that ClpX is required for degradation of the mitochondrial matrix protein DmLRPPRC1, as the substrate-recognition subunit, so the more specific mitochondrial protein catabolic process applies. Proposed replacements: mitochondrial protein catabolic process Supporting Evidence: PMID:28814717 Here, we showed that knockdown of ClpX or ClpP results in an increase of DmLRPPRC1 protein without an increase in the corresponding mRNA (Figs 2 and S4). PMID:28814717 Moreover, we showed that ClpX, which recognize the protein substrates of ClpXP, physically interacts with DmLRPPRC1 in the cells (Fig. |
| GO:0140662 ATP-dependent protein folding chaperone | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: ATP-dependent protein folding chaperone from the ClpX InterPro signature. Reason: ClpX is an ATP-dependent unfoldase that feeds ClpP; a folding-chaperone activity independent of ClpP is documented for some mitochondrial ClpX orthologs but not in flies. Non-core family-level inference. Supporting Evidence: PMID:28814717 ClpXP is composed of a proteolytic subunit, ClpP, and a chaperone-like subunit, ClpX, which carries an ATPase associated with diverse cellular activities (AAA+) domain6, 7. PMID:28814717 Moreover, we showed that ClpX, which recognize the protein substrates of ClpXP, physically interacts with DmLRPPRC1 in the cells (Fig. |
| GO:0141164 mitochondrial protein quality control | TAS PMID:28814717 Drosophila protease ClpXP specifically degrades DmLRPPRC1 co... | ACCEPT | Summary: Author statement that ClpXP functions in mitochondrial protein quality control. Reason: Consistent with conserved ClpXP biology. Supporting Evidence: PMID:28814717 ClpXP is the major protease in the mitochondrial matrix in eukaryotes, and is well conserved among species. |
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Download this section (compressed HTML)Q: Does fly ClpX have ClpP-independent chaperone functions, for example in loading cofactors into mitochondrial enzymes such as ALAS, as reported for yeast and mammalian ClpX?
Q: What degron does ClpX recognize in DmLRPPRC1, and is the C-terminal alanine important?
Experiment: Mutate or extend the C-terminus of DmLRPPRC1 and measure its turnover and co-immunoprecipitation with ClpX in S2 cells expressing catalytically inactive ClpP.
Hypothesis: ClpX recognizes DmLRPPRC1 through a C-terminal degron.
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