mtSSB (lopo, low power) encodes the Drosophila mitochondrial single-stranded DNA-binding protein, a homotetrameric SSB-family protein that coats single-stranded mtDNA exposed during replication. Purified mtSSB binds single-stranded DNA with strong preference over duplex DNA or RNA and stimulates the DNA polymerase and exonuclease activities of DNA polymerase gamma, chiefly by removing template secondary structure and promoting primer recognition and strand initiation. mtSSB is required to maintain mtDNA copy number: lopo mutants and RNAi knockdown deplete mtDNA, lose respiration and arrest cell proliferation, causing late larval lethality.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000262 mitochondrial chromosome | IDA PMID:11294889 Mitochondrial single-stranded DNA-binding protein is require... | ACCEPT | Summary: Mitochondrial chromosome (IDA). Reason: mtSSB coats single-stranded regions of replicating mtDNA within nucleoids. The cached abstract of PMID:11294889 does not describe the localization experiment; the curator's IDA is accepted, consistent with the IBA nucleoid assignment. Supporting Evidence: PMID:11294889 mitochondrial single-stranded DNA-binding protein (mtSSB), which is one of the key components in mtDNA replication and maintenance |
| GO:0003697 single-stranded DNA binding | IBA GO_REF:0000033 | ACCEPT | Summary: Single-stranded DNA binding (IBA). Reason: Core activity of the SSB family. PAINT IBA with WITH/FROM donors including E. coli SSB (P0AGE0), human SSBP1 (Q04837), yeast Rim1 and Drosophila mtSSB itself (FB:FBgn0010438), the latter reflecting its own direct experimental evidence. Supporting Evidence: PMID:7673145 The protein binds to DNA in its single-stranded form with a strong preference over double-stranded DNA or RNA |
| GO:0003697 single-stranded DNA binding | IDA PMID:14754882 Physiological and biochemical defects in functional interact... | ACCEPT | Summary: Single-stranded DNA binding (IDA). Reason: DNA-binding mutants were characterized in this work, which presupposes and measures the ssDNA binding of wild-type mtSSB. Supporting Evidence: PMID:14754882 We show here that DNA-binding mutants of mtSSB are defective in stimulation of DNA synthesis by pol gamma |
| GO:0003697 single-stranded DNA binding | IDA PMID:26446790 Mitochondrial Single-stranded DNA-binding Proteins Stimulate... | ACCEPT | Summary: Single-stranded DNA binding (IDA). Reason: Electron microscopy and biochemistry show that mtSSB binds and opens ssDNA templates, removing secondary structure. Supporting Evidence: PMID:26446790 increasing concentrations of both mtSSBs led to the elimination of template secondary structure and gradual opening of the template DNA |
| GO:0003697 single-stranded DNA binding | IDA PMID:8206370 RNA- and single-stranded DNA-binding (SSB) proteins expresse... | ACCEPT | Summary: Single-stranded DNA binding (IDA). Reason: Purified native and recombinant Drosophila mtSSB binds ssDNA selectively; this is the core molecular function. Supporting Evidence: PMID:8206370 Dm mtSSB specifically binds ss, but not double stranded DNA |
| GO:0003697 single-stranded DNA binding | IEA GO_REF:0000002 | ACCEPT | Summary: Single-stranded DNA binding (IEA/ISS). Reason: Electronic and similarity-based assignments agree with the direct biochemical evidence for ssDNA binding. Supporting Evidence: PMID:7673145 The protein binds to DNA in its single-stranded form with a strong preference over double-stranded DNA or RNA |
| GO:0003697 single-stranded DNA binding | ISS GO_REF:0000110 | ACCEPT | Summary: Single-stranded DNA binding (IEA/ISS). Reason: Electronic and similarity-based assignments agree with the direct biochemical evidence for ssDNA binding. Supporting Evidence: PMID:7673145 The protein binds to DNA in its single-stranded form with a strong preference over double-stranded DNA or RNA |
| GO:0005739 mitochondrion | IDA PMID:7673145 Mitochondrial single-stranded DNA-binding protein from Droso... | ACCEPT | Summary: Mitochondrion. Reason: Native mtSSB was purified from and localized to a mitochondrial fraction. Supporting Evidence: PMID:7673145 its localization to a mitochondrial protein fraction |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: Mitochondrion. Reason: Native mtSSB was purified from and localized to a mitochondrial fraction. Supporting Evidence: PMID:7673145 its localization to a mitochondrial protein fraction |
| GO:0005739 mitochondrion | ISS GO_REF:0000110 | ACCEPT | Summary: Mitochondrion. Reason: Native mtSSB was purified from and localized to a mitochondrial fraction. Supporting Evidence: PMID:7673145 its localization to a mitochondrial protein fraction |
| GO:0006260 DNA replication | IBA GO_REF:0000033 | MODIFY | Summary: DNA replication (IBA/IEA). Reason: Correct but less specific than the evidence supports; replaced by GO:0006264 mitochondrial DNA replication, which the gene already carries by IDA. IBA donors are human SSBP1 (Q04837) and fly mtSSB itself. Proposed replacements: mitochondrial DNA replication |
| GO:0006260 DNA replication | IEA GO_REF:0000002 | MODIFY | Summary: DNA replication (IBA/IEA). Reason: Correct but less specific than the evidence supports; replaced by GO:0006264 mitochondrial DNA replication, which the gene already carries by IDA. IBA donors are human SSBP1 (Q04837) and fly mtSSB itself. Proposed replacements: mitochondrial DNA replication |
| GO:0006264 mitochondrial DNA replication | IDA PMID:10506144 Baculovirus expression reconstitutes Drosophila mitochondria... | ACCEPT | Summary: Mitochondrial DNA replication. Reason: Reconstituted recombinant Pol gamma activity is substantially stimulated by mtSSB, placing mtSSB in the mtDNA replication machinery. Supporting Evidence: PMID:10506144 The recombinant holoenzyme contains DNA polymerase and 3'-5' exonuclease that are stimulated substantially by both salt and mitochondrial single-stranded DNA-binding protein |
| GO:0006264 mitochondrial DNA replication | IDA PMID:11294889 Mitochondrial single-stranded DNA-binding protein is require... | ACCEPT | Summary: Mitochondrial DNA replication. Reason: lopo (mtSSB) mutants and mtSSB RNAi deplete mtDNA, and mtSSB directly stimulates Pol gamma at the replication fork. Supporting Evidence: PMID:11294889 Molecular, histochemical, and physiological experiments show a drastic decrease in mtDNA content PMID:14754882 Overexpression of mtSSB restores cell growth rate and the copy number of mtDNA |
| GO:0006264 mitochondrial DNA replication | IDA PMID:26446790 Mitochondrial Single-stranded DNA-binding Proteins Stimulate... | ACCEPT | Summary: Mitochondrial DNA replication. Reason: Reconstitution with purified Pol gamma shows mtSSB acting as a replisome component that stimulates DNA synthesis. Supporting Evidence: PMID:26446790 The activity of the mitochondrial replicase, DNA polymerase Ξ³ (Pol Ξ³) is stimulated by another key component of the mitochondrial replisome, the mitochondrial single-stranded DNA-binding protein (mtSSB) |
| GO:0008047 enzyme activator activity | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: Enzyme activator activity (IBA). Reason: IBA seeded only by E. coli SSB (P0AGE0). mtSSB does stimulate Pol gamma, but the stimulation has been traced mainly to organization of the template DNA rather than to binding and allosterically activating the enzyme, which is what GO:0008047 implies. The stimulation is better captured by the process term GO:1900264 positive regulation of DNA-directed DNA polymerase activity. Supporting Evidence: PMID:26446790 We conclude that organization of the template DNA is the major factor contributing to the stimulation of Pol Ξ³ activity |
| GO:0042645 mitochondrial nucleoid | IBA GO_REF:0000033 | ACCEPT | Summary: Mitochondrial nucleoid (IBA). Reason: IBA from the mtSSB clade (human SSBP1 Q04837, yeast Rim1 SGD:S000007222, fly mtSSB). mtSSB acts on nucleoid-associated replicating mtDNA; nucleoid is the most informative location. |
| GO:0090297 positive regulation of mitochondrial DNA replication | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Positive regulation of mitochondrial DNA replication. Reason: mtSSB levels positively control mtDNA copy number (RNAi depletes mtDNA, re-expression restores it), which is what this term records. Mechanistically mtSSB is a structural component of the replisome rather than a regulator; its core process is captured by GO:0006264 mitochondrial DNA replication. Supporting Evidence: PMID:14754882 Overexpression of mtSSB restores cell growth rate and the copy number of mtDNA |
| GO:0090297 positive regulation of mitochondrial DNA replication | IMP PMID:14754882 Physiological and biochemical defects in functional interact... | KEEP AS NON CORE | Summary: Positive regulation of mitochondrial DNA replication. Reason: mtSSB levels positively control mtDNA copy number (RNAi depletes mtDNA, re-expression restores it), which is what this term records. Mechanistically mtSSB is a structural component of the replisome rather than a regulator; its core process is captured by GO:0006264 mitochondrial DNA replication. Supporting Evidence: PMID:14754882 Overexpression of mtSSB restores cell growth rate and the copy number of mtDNA |
| GO:0090592 DNA synthesis involved in DNA replication | IDA PMID:10329675 Functional interactions of mitochondrial DNA polymerase and ... | MODIFY | Summary: DNA synthesis involved in DNA replication. Reason: mtSSB stimulates primer recognition and strand initiation by Pol gamma, so it participates in replicative DNA synthesis; the term does not capture that this is mtDNA replication, so it is replaced by GO:0006264 mitochondrial DNA replication, which the gene already carries. Proposed replacements: mitochondrial DNA replication Supporting Evidence: PMID:10329675 mtSSB enhances primer recognition and binding and stimulates 30-fold the rate of initiation of DNA strands |
| GO:0090592 DNA synthesis involved in DNA replication | IDA PMID:15537632 Mutations in the spacer region of Drosophila mitochondrial D... | MODIFY | Summary: DNA synthesis involved in DNA replication. Reason: Pol gamma spacer mutants were analyzed for their functional interaction with mtSSB in DNA synthesis; correct but less specific than GO:0006264 mitochondrial DNA replication, which the gene already carries. Proposed replacements: mitochondrial DNA replication Supporting Evidence: PMID:15537632 several mutations affect differentially DNA polymerase and exonuclease activity and/or functional interactions with mitochondrial single-stranded DNA-binding protein |
| GO:1900264 positive regulation of DNA-directed DNA polymerase activity | IMP PMID:14754882 Physiological and biochemical defects in functional interact... | ACCEPT | Summary: Positive regulation of DNA-directed DNA polymerase activity. Reason: mtSSB stimulates the polymerase (and exonuclease) activity of Pol gamma, and DNA-binding mutants of mtSSB lose this stimulation; this is a defining property of mtSSB in the replisome. Supporting Evidence: PMID:14754882 We show here that DNA-binding mutants of mtSSB are defective in stimulation of DNA synthesis by pol gamma |
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Experiment: Reconstitute the Drosophila minimal mitochondrial replisome (Pol gamma, mtDNA helicase, mtSSB) and measure rolling-circle synthesis with wild-type versus DNA-binding-defective mtSSB.
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