The yeast RAVE (Regulator of H+-ATPase of Vacuolar and Endosomal membranes) complex and Rabconnectin-3 complexes of higher eukaryotes regulate acidification of organelles such as lysosomes and endosomes by catalyzing V-ATPase assembly. V-ATPases are highly conserved proton pumps consisting of a peripheral V1 subcomplex that contains the sites of ATP hydrolysis, attached to an integral membrane Vo subcomplex that forms the transmembrane proton pore. Reversible disassembly of the V-ATPase is a conserved regulatory mechanism that occurs in response to multiple signals, serving to tune ATPase activity and compartment acidification to changing extracellular conditions. Signals such as glucose deprivation can induce release of V1 from Vo, which inhibits both ATPase activity and proton transport. Reassembly of V1 with Vo restores ATP-driven proton transport, but requires assistance of the RAVE or Rabconnectin-3 complexes. Glucose deprivation triggers V-ATPase disassembly in yeast and is accompanied by binding of RAVE to V1 subcomplexes. Upon glucose readdition, RAVE catalyzes both recruitment of V1 to the vacuolar membrane and its reassembly with Vo. The RAVE complex can be recruited to the vacuolar membrane by glucose in the absence of V1 subunits, indicating that the interaction between RAVE and the Vo membrane domain is glucose-sensitive. Yeast RAVE complexes also distinguish between organelle-specific isoforms of the Vo a-subunit and thus regulate distinct V-ATPase subpopulations. Rabconnectin-3 complexes in higher eukaryotes appear to be functionally equivalent to yeast RAVE. Originally isolated as a two-subunit complex from rat brain, the Rabconnectin-3 complex has regions of homology with yeast RAVE and was shown to interact with V-ATPase subunits and promote endosomal acidification. Current understanding of the structure and function of RAVE and Rabconnectin-3 complexes, their interactions with the V-ATPase, their role in signal-dependent modulation of organelle acidification, and their impact on downstream pathways will be discussed.
Mutations in the human Rogdi protein cause Kohlschutter‐Tonz syndrome, which is characterized by early‐onset seizures, developmental defects, and defective deposition of tooth enamel (amelogenesis imperfecta). The cellular function of Rogdi is not known. The yeast RAVE (regulator of ATPase of vacuoles and endosomes) complex and mammalian Rabconnectin‐3 complexes catalyze assembly of certain subpopulations of the V‐ATPase proton pump and thus help to regulate pH homeostasis. The yeast RAVE complex is composed of three subunits: Rav1, Rav2, and Skp1. Skp1 is a multifunctional scaffold protein, but both Rav1 and Rav2 act specifically in V‐ATPase assembly. Rabconnectin‐3 complexes have been reported to contain two larger subunits, Rbcn3α and 3β; both subunits are predicted to have structural similarity to Rav1. No mammalian homologue of yeast Rav2 had been identified. However, despite limited sequence homology, we find that the yeast Rav2 sequence can be modeled with very high confidence on a recent structure of human Rogdi (H. Lee et al. (2017) Sci. Rep. 7:3972). Expression of human Rogdi in yeast complements the V‐ATPase‐associated growth defects of a yeast rav2∆ mutant. Complementation requires the presence of Rav1, suggesting that Rogdi acts as part of the RAVE complex rather than bypassing RAVE function. Consistent with this, Rogdi copurifies with a FLAG‐tagged Rav1 protein. Yeast Rav2 binds to the N‐terminal β‐propeller region of Rav1. Two‐hybrid assays indicate that Rogdi can interact with the same region of Rav1, as well as binding to the N‐terminal β‐propeller regions of Rbcn3α and 3β isoforms. Based on these data, we hypothesize that Rogdi is a subunit of mammalian Rabconnectin‐3 complexes. WDR72 is a possible Rbcn3β isoform in humans. Mutations in the N‐terminal β‐propeller domain of WDR72 also cause amelogenesis imperfecta, as well as distal renal tubule acidosis, a disease associated with defects in V‐ATPase activity. We are testing whether these mutations affect the interaction between Rogdi and WDR72, potentially linking Rogdi with both Rabconnectin‐3 and V‐ATPase function.
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