biostudies-literatureUnknownKhan MMNIGMS NIH HHSThe eukaryotic vacuolar H⁺-ATPase (V-ATPase) is regulated by reversible disassembly into autoinhibited V₁-ATPase and V_o proton channel subcomplexes, a mode of regulation conserved from yeast to humans. While signals that govern V-ATPase assembly have been studied in the cellular context, the molecular mechanisms of the process at the level of the enzyme remain poorly understood. We recently discovered that Oxr1p, one of the two TLDc domain proteins in yeast, is essential for rapid V-ATPase <i>disassembly in vivo</i>. How the second TLDc protein, Rtc5p, functions in reversible disassembly, however, is less clear. Here we find that Rtc5p promotes assembly of functional holo V-ATPase from purified V₁ and V_o subcomplexes <i>in vitro</i>. CbioRxiv : the preprint server for biology2025.05.24.655954https://www.ebi.ac.uk/biostudies/studies/S-EPMC12139955biostudies-literatureInteraction of yeast V-ATPase with TLDc protein Rtc5p.PMC12139955R35 GM141908Khan MMEbrahimi ROot RAWilkens SfalseInteraction of yeast V-ATPase with TLDc protein Rtc5p.The eukaryotic vacuolar H⁺-ATPase (V-ATPase) is regulated by reversible disassembly into autoinhibited V₁-ATPase and V_o proton channel subcomplexes, a mode of regulation conserved from yeast to humans. While signals that govern V-ATPase assembly have been studied in the cellular context, the molecular mechanisms of the process at the level of the enzyme remain poorly understood. We recently discovered that Oxr1p, one of the two TLDc domain proteins in yeast, is essential for rapid V-ATPase <i>disassembly in vivo</i>. How the second TLDc protein, Rtc5p, functions in reversible disassembly, however, is less clear. Here we find that Rtc5p promotes assembly of functional holo V-ATPase from purified V₁ and V_o subcomplexes <i>in vitro</i>. C2025-01-01T00:00:00Z2025 May2026-04-26T03:15:22.164Z2026-04-26T03:10:42.686ZS-EPMC121399554047562010.1101/2025.05.24.655954