The role of the COOH terminus of Sec2p in the transport of post-Golgi vesicles.
Sec2p is required for the polarized transport of secretory vesicles in S. cerevisiae. The Sec2p NH(2) terminus encodes an exchange factor for the Rab protein Sec4p. Sec2p associates with vesicles and in Sec2p COOH-terminal mutants Sec4p and vesicles no longer accumulate at bud tips. Thus, the Sec2p COOH terminus functions in targeting vesicles, however, the mechanism of function is unknown. We found comparable exchange activity for truncated and full-length Sec2 proteins, implying that the COOH terminus does not alter the exchange rate. Full-length Sec2-GFP, similar to Sec4p, concentrates at bud tips. A COOH-terminal 58-amino acid domain is necessary but not sufficient for localization. Sec2p localization depends on actin, Myo2p and Sec1p, Sec6p, and Sec9p function. Full-length, but not COOH-terminally truncated Sec2 proteins are enriched on membranes. Membrane association of full-length Sec2p is reduced in sec6-4 and sec9-4 backgrounds at 37 degrees C but unaffected at 25 degrees C. Taken together, these data correlate loss of localization of Sec2 proteins with reduced membrane association. In addition, Sec2p membrane attachment is substantially Sec4p independent, supporting the notion that Sec2p interacts with membranes via an unidentified Sec2p receptor, which would increase the accessibility of Sec2p exchange activity for Sec4p.
Elkind
N B
NB
Department of Cell Biology, Yale University, New Haven, Connecticut, 06510, USA.
Walch-Solimena
C
C
Novick
P J
PJ
D000199:Actins / Q000378:metabolism; D001692:Biological Transport; D002455:Cell Division; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D016764:Cell Polarity; D003594:Cytoplasmic Granules / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal / Q000235:genetics / Q000502:physiology; D006056:Golgi Apparatus / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors; D007700:Kinetics; D010766:Phosphorylation; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion / Q000235:genetics; D013696:Temperature; D014617:Vacuoles / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism
D000199:Actins; D005656:Fungal Proteins; D020662:Guanine Nucleotide Exchange Factors; D011993:Recombinant Fusion Proteins; C064456:SEC2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D019204:GTP-Binding Proteins; C064320:SEC4 protein, S cerevisiae; D020691:rab GTP-Binding Proteins