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. 1998 Jun;149(2):833–841. doi: 10.1093/genetics/149.2.833

A high copy suppressor screen reveals genetic interactions between BET3 and a new gene. Evidence for a novel complex in ER-to-Golgi transport.

Y Jiang 1, A Scarpa 1, L Zhang 1, S Stone 1, E Feliciano 1, S Ferro-Novick 1
PMCID: PMC1460158  PMID: 9611195

Abstract

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.

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Selected References

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  1. Barlowe C. Coupled ER to Golgi transport reconstituted with purified cytosolic proteins. J Cell Biol. 1997 Dec 1;139(5):1097–1108. doi: 10.1083/jcb.139.5.1097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Carlson M., Botstein D. Two differentially regulated mRNAs with different 5' ends encode secreted with intracellular forms of yeast invertase. Cell. 1982 Jan;28(1):145–154. doi: 10.1016/0092-8674(82)90384-1. [DOI] [PubMed] [Google Scholar]
  3. Chapman R. E., Munro S. The functioning of the yeast Golgi apparatus requires an ER protein encoded by ANP1, a member of a new family of genes affecting the secretory pathway. EMBO J. 1994 Oct 17;13(20):4896–4907. doi: 10.1002/j.1460-2075.1994.tb06817.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Collins R. N., Brennwald P., Garrett M., Lauring A., Novick P. Interactions of nucleotide release factor Dss4p with Sec4p in the post-Golgi secretory pathway of yeast. J Biol Chem. 1997 Jul 18;272(29):18281–18289. doi: 10.1074/jbc.272.29.18281. [DOI] [PubMed] [Google Scholar]
  5. Ferro-Novick S., Jahn R. Vesicle fusion from yeast to man. Nature. 1994 Jul 21;370(6486):191–193. doi: 10.1038/370191a0. [DOI] [PubMed] [Google Scholar]
  6. Griff I. C., Schekman R., Rothman J. E., Kaiser C. A. The yeast SEC17 gene product is functionally equivalent to mammalian alpha-SNAP protein. J Biol Chem. 1992 Jun 15;267(17):12106–12115. [PubMed] [Google Scholar]
  7. Hardwick K. G., Pelham H. R. SED5 encodes a 39-kD integral membrane protein required for vesicular transport between the ER and the Golgi complex. J Cell Biol. 1992 Nov;119(3):513–521. doi: 10.1083/jcb.119.3.513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  9. Lian J. P., Ferro-Novick S. Bos1p, an integral membrane protein of the endoplasmic reticulum to Golgi transport vesicles, is required for their fusion competence. Cell. 1993 May 21;73(4):735–745. doi: 10.1016/0092-8674(93)90253-m. [DOI] [PubMed] [Google Scholar]
  10. Lian J. P., Stone S., Jiang Y., Lyons P., Ferro-Novick S. Ypt1p implicated in v-SNARE activation. Nature. 1994 Dec 15;372(6507):698–701. doi: 10.1038/372698a0. [DOI] [PubMed] [Google Scholar]
  11. Louvard D., Reggio H., Warren G. Antibodies to the Golgi complex and the rough endoplasmic reticulum. J Cell Biol. 1982 Jan;92(1):92–107. doi: 10.1083/jcb.92.1.92. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Mayer A., Wickner W., Haas A. Sec18p (NSF)-driven release of Sec17p (alpha-SNAP) can precede docking and fusion of yeast vacuoles. Cell. 1996 Apr 5;85(1):83–94. doi: 10.1016/s0092-8674(00)81084-3. [DOI] [PubMed] [Google Scholar]
  13. Muhlrad D., Hunter R., Parker R. A rapid method for localized mutagenesis of yeast genes. Yeast. 1992 Feb;8(2):79–82. doi: 10.1002/yea.320080202. [DOI] [PubMed] [Google Scholar]
  14. Rossi G., Kolstad K., Stone S., Palluault F., Ferro-Novick S. BET3 encodes a novel hydrophilic protein that acts in conjunction with yeast SNAREs. Mol Biol Cell. 1995 Dec;6(12):1769–1780. doi: 10.1091/mbc.6.12.1769. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ruohola H., Kabcenell A. K., Ferro-Novick S. Reconstitution of protein transport from the endoplasmic reticulum to the Golgi complex in yeast: the acceptor Golgi compartment is defective in the sec23 mutant. J Cell Biol. 1988 Oct;107(4):1465–1476. doi: 10.1083/jcb.107.4.1465. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sacher M., Jiang Y., Barrowman J., Scarpa A., Burston J., Zhang L., Schieltz D., Yates J. R., 3rd, Abeliovich H., Ferro-Novick S. TRAPP, a highly conserved novel complex on the cis-Golgi that mediates vesicle docking and fusion. EMBO J. 1998 May 1;17(9):2494–2503. doi: 10.1093/emboj/17.9.2494. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Sacher M., Stone S., Ferro-Novick S. The synaptobrevin-related domains of Bos1p and Sec22p bind to the syntaxin-like region of Sed5p. J Biol Chem. 1997 Jul 4;272(27):17134–17138. doi: 10.1074/jbc.272.27.17134. [DOI] [PubMed] [Google Scholar]
  18. Sapperstein S. K., Lupashin V. V., Schmitt H. D., Waters M. G. Assembly of the ER to Golgi SNARE complex requires Uso1p. J Cell Biol. 1996 Mar;132(5):755–767. doi: 10.1083/jcb.132.5.755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Shim J., Newman A. P., Ferro-Novick S. The BOS1 gene encodes an essential 27-kD putative membrane protein that is required for vesicular transport from the ER to the Golgi complex in yeast. J Cell Biol. 1991 Apr;113(1):55–64. doi: 10.1083/jcb.113.1.55. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sikorski R. S., Hieter P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics. 1989 May;122(1):19–27. doi: 10.1093/genetics/122.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Stone S., Sacher M., Mao Y., Carr C., Lyons P., Quinn A. M., Ferro-Novick S. Bet1p activates the v-SNARE Bos1p. Mol Biol Cell. 1997 Jul;8(7):1175–1181. doi: 10.1091/mbc.8.7.1175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Söllner T., Whiteheart S. W., Brunner M., Erdjument-Bromage H., Geromanos S., Tempst P., Rothman J. E. SNAP receptors implicated in vesicle targeting and fusion. Nature. 1993 Mar 25;362(6418):318–324. doi: 10.1038/362318a0. [DOI] [PubMed] [Google Scholar]
  23. Wilson D. W., Wilcox C. A., Flynn G. C., Chen E., Kuang W. J., Henzel W. J., Block M. R., Ullrich A., Rothman J. E. A fusion protein required for vesicle-mediated transport in both mammalian cells and yeast. Nature. 1989 Jun 1;339(6223):355–359. doi: 10.1038/339355a0. [DOI] [PubMed] [Google Scholar]
  24. von Mollard G. F., Nothwehr S. F., Stevens T. H. The yeast v-SNARE Vti1p mediates two vesicle transport pathways through interactions with the t-SNAREs Sed5p and Pep12p. J Cell Biol. 1997 Jun 30;137(7):1511–1524. doi: 10.1083/jcb.137.7.1511. [DOI] [PMC free article] [PubMed] [Google Scholar]

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