Molecular mechanisms of disassembly and reassembly of the V₁ and V₀ domains of the yeast vacuolar H⁺-ATPase

Vacuolar H⁺-ATPases acidify intracellular mpartments by catalyzing the hydrolysis of cytosolic ATP coupled to proton translocation across the organelle membrane. The enzyme is a multisubunit complex organized into two domains, V₁ and V_o. V₁ is peripherally attached to the cytosolic side of the membrane and contains the catalytic sites for ATP hydrolysis. V_o forms a transmembranous proton channel and anchors V₁. It has been proposed that reversible association between V₁ and V_o is a mechanism for regulation of organelle acidification in vivo. We studied the molecular triggers of in vivo disassembly and reassembly of V₁ and V_o in yeast. V₁ and V_o interactions can be controlled by extracellular nutrients. The pool of V₁V_o complexes is modulated by the amount of glucose present in the medium on both short-term and long-term basis. Association between V₁ and V_o may be regulated by the glycolytic flux. First, reassembly of V₁ and V_o is induced only by addition of glucose, fructose or mannose, three rapidly metabolized sugars, to glucose-deprived cells. Second, accumulation of glucose-6-phosphate alone cannot maintain enzyme assembly. Third, disassembly naturally occurs when exponentially growing cells deplete glucose and reach stationary phase. The effect of glycolytic flux on the vacuolar H⁺-ATPase is likely to be indirect. A transient decrease in ATP concentration with glucose deprivation occurs on a sufficiently rapid time scale to help trigger disassembly, but increases in cellular ATP concentrations with glucose readdition cannot account for reassembly. When ATP levels are low, catalysis may induce a structural conformation that favors dissociation of V₁ from V_o. Accordingly, impairment of activity significantly reduces disassembly. A drop in cytosolic pH with initiation of glycolysis, could provide a signal and a reason for reassembly.