Topology control by a conserved cysteine pair in the OMM-protein CCDC127 enables MICOS interaction

Riemer, J.; Zarges, C.; Schepsky, P.; Rothemann, R. A.; Bock-Bierbaum, T.; von der Malsburg, A.; Baumann, L.; Trifunovic, A.; Daumke, O.; van der Laan, M.; von der Malsburg, K.

doi:10.64898/2025.12.12.693940

bioRxiv · 10.64898/2025.12.12.693940

Topology control by a conserved cysteine pair in the OMM-protein CCDC127 enables MICOS interaction

Riemer, J.·Zarges, C.·Schepsky, P.·Rothemann, R. A.·Bock-Bierbaum, T.·von der Malsburg, A.·Baumann, L.·Trifunovic, A.·Daumke, O.·van der Laan, M.·von der Malsburg, K.

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Abstract

Mitochondrial disulfide relay substrates beyond the canonical substrates remain incompletely defined. Revisiting the human MIA40 interactome with enhanced depth, we identified CCDC127 as a previously unrecognized substrate candidate. CCDC127 contains a single transmembrane segment and a conserved C-terminal helical bundle domain (CHB). Comprehensive proteomic and biochemical analyses revealed that, contrary to earlier reports, CCDC127 adopts an Nout-Cin topology in the outer mitochondrial membrane (OMM) with its CHB residing in the intermembrane space (IMS). CCDC127 undergoes oxidation by the disulfide relay, forming a long-range intramolecular disulfide bond between C174 and C219. Loss of these cysteines disrupts correct OMM insertion, inverts transmembrane topology and triggers proteasome-dependent degradation, establishing the disulfide as a key determinant of CCDC127 maturation. Interactome analyses identified MICOS components--particularly the MIC60/MIC19 module--as major partner proteins required for the stability of large oligomeric CCDC127 complexes. CCDC127 deficiency impaired cellular proliferation, influenced phospholipid levels, and caused grossly altered cristae morphology. Together, CCDC127 emerges as a MICOS-associated OMM protein essential for mitochondrial membrane organization and lipid homeostasis.

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Riemer, J., Zarges, C., Schepsky, P., Rothemann, R. A., Bock-Bierbaum, T., von der Malsburg, A., Baumann, L., Trifunovic, A., Daumke, O., van der Laan, M., von der Malsburg, K.. 2025-12-15. Topology control by a conserved cysteine pair in the OMM-protein CCDC127 enables MICOS interaction. https://doi.org/10.64898/2025.12.12.693940

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