Two GTPase isoforms, Ypt31p and Ypt32p, are essential for Golgi function in yeast.
In eukaryotic cells, monomeric GTPases of the Ypt/Rab family function as regulators at defined steps of vesicular transport in exo- and endocytosis. Here we report on the isolation and characterization of two genes (YPT31 and YPT32) of the yeast Saccharomyces cerevisiae which encode members of the Ypt family exhibiting >80% sequence identity. Whereas the disruption of one of the two genes was phenotypically neutral, the disruption of both YPT31 and YPT32 led to lethality. Depletion of wild-type Ypt31p or of a short-lived ubiquitin-Ypt31p in a ypt32 null background led to a massive accumulation of Golgi-like membranes, an inhibition of invertase secretion and defects in vacuolar protein maturation. Similar alterations were observed in a conditional-lethal ypt31-1 mutant at 30 min after shift to the non-permissive temperature. According to subcellular fractionation, a significant part of Ypt31p appeared to be located in Golgi-enriched membrane fractions. In accordance with this, indirect immunofluorescence using affinity-purified anti-Ypt31p antibodies gave a punctate staining similar to that observed with Golgi-located proteins. From the phenotypic alterations observed in ypt31 and ypt32 mutants, it seems likely that the two GTPases are involved in intra-Golgi transport or in the formation of transport vesicles at the most distal Golgi compartment.
Benli
M
M
Department of Molecular Genetics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
Döring
F
F
Robinson
D G
DG
Yang
X
X
Gallwitz
D
D
D000595:Amino Acid Sequence; D000818:Animals; D004285:Dogs; D020558:GTP Phosphohydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D006026:Glycoside Hydrolases / Q000378:metabolism; D006056:Golgi Apparatus / Q000502:physiology* / Q000648:ultrastructure; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D005810:Multigene Family; D016296:Mutagenesis; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000648:ultrastructure; D017386:Sequence Homology, Amino Acid; D043324:beta-Fructofuranosidase
D007527:Isoenzymes; D011994:Recombinant Proteins; D006026:Glycoside Hydrolases; D043324:beta-Fructofuranosidase; D020558:GTP Phosphohydrolases