Sec34p, a protein required for vesicle tethering to the yeast Golgi apparatus, is in a complex with Sec35p.
A screen for mutants of Saccharomyces cerevisiae secretory pathway components previously yielded sec34, a mutant that accumulates numerous vesicles and fails to transport proteins from the ER to the Golgi complex at the restrictive temperature (Wuestehube, L.J., R. Duden, A. Eun, S. Hamamoto, P. Korn, R. Ram, and R. Schekman. 1996. Genetics. 142:393-406). We find that SEC34 encodes a novel protein of 93-kD, peripherally associated with membranes. The temperature-sensitive phenotype of sec34-2 is suppressed by the rab GTPase Ypt1p that functions early in the secretory pathway, or by the dominant form of the ER to Golgi complex target-SNARE (soluble N-ethylmaleimide sensitive fusion protein attachment protein receptor)-associated protein Sly1p, Sly1-20p. Weaker suppression is evident upon overexpression of genes encoding the vesicle tethering factor Uso1p or the vesicle-SNAREs Sec22p, Bet1p, or Ykt6p. This genetic suppression profile is similar to that of sec35-1, a mutant allele of a gene encoding an ER to Golgi vesicle tethering factor and, like Sec35p, Sec34p is required in vitro for vesicle tethering. sec34-2 and sec35-1 display a synthetic lethal interaction, a genetic result explained by the finding that Sec34p and Sec35p can interact by two-hybrid analysis. Fractionation of yeast cytosol indicates that Sec34p and Sec35p exist in an approximately 750-kD protein complex. Finally, we describe RUD3, a novel gene identified through a genetic screen for multicopy suppressors of a mutation in USO1, which suppresses the sec34-2 mutation as well.
VanRheenen
S M
SM
Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA.
Cao
X
X
Sapperstein
S K
SK
Chiang
E C
EC
Lupashin
V V
VV
Barlowe
C
C
Waters
M G
MG
D033942:Adaptor Proteins, Vesicular Transport; D000595:Amino Acid Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D002458:Cell Fractionation; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017353:Gene Deletion; D005838:Genotype; D006056:Golgi Apparatus / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D010957:Plasmids; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins
D033942:Adaptor Proteins, Vesicular Transport; C112749:COG2 protein, S cerevisiae; C401396:COG3 protein, S cerevisiae; D002352:Carrier Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; C400547:RUD3 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C086888:USO1 protein, S cerevisiae; D033921:Vesicular Transport Proteins