title: Tracking of the dynamic localization of the Rab-specific HOPS subunits reveal their distinct interaction with Ypt7 and vacuoles.
abstract: Endosomal and vacuole fusion depends on the two homologous tethering complexes CORVET and HOPS. HOPS binds the activated Rab GTPase Ypt7 via two distinct subunits, Vps39 and Vps41. To understand the participation and possible polarity of Vps41 and Vps39 during tethering, we used an in vivo approach. For this, we established the ligand-induced relocalization to the plasma membrane, using the Mon1-Ccz1 GEF complex that activates Ypt7 on endosomes. We then employed slight overexpression to compare the mobility of the HOPS-specific Vps41 and Vps39 subunits during this process. Our data indicate an asymmetry in the Rab-specific interaction of the two HOPS subunits: Vps39 is more tightly bound to the vacuole, and relocalizes the entire vacuole to the plasma membrane, whereas Vps41 behaved like the more mobile subunit. This is due to their specific Rab binding, as the mobility of both subunits was similar inypt7∆ cells. In contrast, both HOPS subunits were far less mobile if tagged endogenously, suggesting that the entire HOPS complex is tightly bound to the vacuole in vivo. Similar results were obtained for the endosomal association of CORVET, when we followed its Rab-specific subunit Vps8. Our data provide in vivo evidence for distinct Rab specificity within HOPS, which may explain its function during tethering, and indicate that these tethering complexes are less mobile within the cell than previously anticipated.
authors.affiliation: Biochemistry section; Department of Biology/Chemistry; University of Osnabrück; Osnabrück, Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Auffarth
authors.affiliation: Biochemistry section; Department of Biology/Chemistry; University of Osnabrück; Osnabrück, Germany.
authors.forename: Henning
authors.initials: H
authors.lastname: Arlt
authors.affiliation: Biochemistry section; Department of Biology/Chemistry; University of Osnabrück; Osnabrück, Germany.
authors.forename: Jens
authors.initials: J
authors.lastname: Lachmann
authors.affiliation: Biochemistry section; Department of Biology/Chemistry; University of Osnabrück; Osnabrück, Germany.
authors.forename: Margarita
authors.initials: M
authors.lastname: Cabrera
authors.affiliation: Biochemistry section; Department of Biology/Chemistry; University of Osnabrück; Osnabrück, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Ungermann
keywords: HOPS; Vps39; Vps41; endosome; tethering complex; vacuole