Abstract
VPS9 domains can act as guanosine nucleotide exchange factors (GEFs) against small G proteins of the Rab5 family. Saccharomyces cerevisiae vps9Δ mutants have trafficking defects considerably less severe than multiple deletions of the three cognate Rab5 paralogs (Vps21, Ypt52, and Ypt53). Here, we show that Muk1, which also contains a VPS9 domain, acts as a second GEF against Vps21, Ypt52, and Ypt53. Muk1 is partially redundant with Vps9 in vivo, with vps9Δ muk1Δ double mutant cells displaying hypersensitivity to temperature and ionic stress, as well as profound impairments in endocytic and Golgi endosome trafficking, including defects in sorting through the multivesicular body. Cells lacking both Vps9 and Muk1 closely phenocopy double and triple knock-out strains lacking Rab5 paralogs. Microscopy and overexpression experiments demonstrate that Vps9 and Muk1 have distinct localization determinants. These experiments establish Muk1 as the second Rab5 GEF in budding yeast.
Highlights
Vps9 is a guanine nucleotide exchange factor (GEF) that activates Rab5 paralogs in yeast
Using biochemical and genetic analyses, we show that Muk1 is a specific GEF for yeast Rab5 proteins and that its in vivo function is partially redundant with the major GEF of the Rab5 family, Vps9
Growth Phenotypes of Muk1 Deletion and Overexpression in Cells Lacking Vps9—The three Rab5 paralogs of budding yeast (Vps21, Ypt52, and Ypt53) are partially redundant, with single mutants of Vps21 displaying clear sorting and morphology phenotypes that are further enhanced by deletion of Ypt52, Ypt53, or both [3, 4, 6]
Summary
Vps is a guanine nucleotide exchange factor (GEF) that activates Rab paralogs in yeast. VPS9 domains can act as guanosine nucleotide exchange factors (GEFs) against small G proteins of the Rab family. Muk is partially redundant with Vps in vivo, with vps9⌬ muk1⌬ double mutant cells displaying hypersensitivity to temperature and ionic stress, as well as profound impairments in endocytic and Golgi endosome trafficking, including defects in sorting through the multivesicular body. Cells lacking both Vps and Muk closely phenocopy double and triple knock-out strains lacking Rab paralogs. The mammalian Rin proteins (Rin1–3) contain VPS9 domains and act as GEFs against Rab isoforms, suggesting that VPS9 domains generally function to stimulate signaling by members of the Rab family [15,16,17]. Using biochemical and genetic analyses, we show that Muk is a specific GEF for yeast Rab proteins and that its in vivo function is partially redundant with the major GEF of the Rab family, Vps
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