A high copy suppressor screen reveals genetic interactions between BET3 and a new gene. Evidence for a novel complex in ER-to-Golgi transport.
The BET3 gene in the yeast Saccharomyces cerevisiae encodes a 22-kD hydrophilic protein that is required for vesicular transport between the ER and Golgi complex. To gain insight into the role of Bet3p, we screened for genes that suppress the growth defect of the temperature-sensitive bet3 mutant at 34 degrees. This high copy suppressor screen resulted in the isolation of a new gene, called BET5. BET5 encodes an essential 18-kD hydrophilic protein that in high copy allows growth of the bet3-1 mutant, but not other ER accumulating mutants. This strong and specific suppression is consistent with the fact that Bet3p and Bet5p are members of the same complex. Using PCR mutagenesis, we generated a temperature-sensitive mutation in BET5 (bet5-1) that blocks the transport of carboxypeptidase Y to the vacuole and prevents secretion of the yeast pheromone alpha-factor at 37 degrees. The precursor forms of these proteins that accumulate in this mutant are indicative of a block in membrane traffic between the ER and Golgi apparatus. High copy suppressors of the bet5-1 mutant include several genes whose products are required for ER-to-Golgi transport (BET1, SEC22, USO1 and DSS4) and the maintenance of the Golgi (ANP1). These findings support the hypothesis that Bet5p acts in conjunction with Bet3p to mediate a late stage in ER-to-Golgi transport. The identification of mammalian homologues of Bet3p and Bet5p implies that the Bet3p/Bet5p complex is highly conserved in evolution.
Jiang
Y
Y
Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
Scarpa
A
A
Zhang
L
L
Stone
S
S
Feliciano
E
E
Ferro-Novick
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport / Q000235:genetics; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D017124:Conserved Sequence; D004721:Endoplasmic Reticulum / Q000235:genetics* / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D018628:Gene Dosage; D005800:Genes, Fungal; D006056:Golgi Apparatus / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011233:Precipitin Tests; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013489:Suppression, Genetic; D033921:Vesicular Transport Proteins
C098389:BET3 protein, S cerevisiae; C114038:BET5 protein, S cerevisiae; D002352:Carrier Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins