In Saccharomyces cerevisiae, the YPT6 gene encodes the homologue of the mammalian Rab6 protein found in the Golgi apparatus. Deletion of YPT6 in yeast produces a phenotype showing temperature-sensitive growth and partial missorting of the vacuolar enzyme, carboxypeptidase Y. To identify proteins that might: (1) interact with Ypt6p; or (2) act in the same pathway, we have isolated four multicopy suppressors, named SYS1, SYS2, SYS3 and SYS5, that can complement the temperature-sensitive growth phenotype of the ypt6 null mutant. On high expression, these genes are also able to partially suppress the missorting of carboxypeptidase Y.SYS2 on a multicopy plasmid suppresses in addition the temperature-sensitive phenotype of sec7-1, a mutant defective in transport between and from the Golgi compartment. Gene disruption of SYS1 and SYS2 did not result in significant growth defects. However, deletion of SYS1 and/or SYS2 in the ypt6 null mutant enhances defects in vacuolar protein sorting and in cell growth. Whereas protein secretion was not significantly affected in these mutants, the processing of alpha-factor precursor by the Kex2 protease was inhibited, suggesting a function of YPT6 and its null mutant suppressors in transport between the late Golgi and a prevacuolar, endosome-like compartment.

Aalto
M. K.
,
Ronne
H.
,
Keränen
S.
(
1993
).
Yeast syntaxins Sso1p and Sso2p belong to a family of related membrane proteins that function in vesicular transport.
EMBO J
12
,
4095
4104
Antony
C.
,
Cibert
C.
,
Geraud
G.
,
Santa Maria
A.
,
Maro
B.
,
Mayau
V.
,
Goud
B.
(
1992
).
The small GTP-binding protein rab6p is distributed from medial Golgi to the trans -Golgi network as determined by a confocal microscopic approach.
J. Cell Sci
103
,
785
796
Banfield
D. K.
,
Lewis
M. J.
,
Pelham
H. R. B.
(
1995
).
A SNARE-like protein required for traffic through the Golgi complex.
Nature
375
,
806
809
Bankaitis
V. A.
,
Malehorn
D. E.
,
Emr
S. D.
,
Greene
R.
(
1989
).
The Saccharomyces cerevisiaeSEC14 gene encodes a cytosolic factor that is required for transport of secretory proteins from the yeast Golgi complex.
J. Cell Biol
108
,
1271
1281
Bankaitis
V. A.
,
Aitken
J. R.
,
Cleves
A. E.
,
Dowhan
W.
(
1990
).
An essential role for a phospholipid transfer protein in yeast Golgi function.
Nature
347
,
561
562
Botstein
D.
,
Falco
S. C.
,
Stewart
S. E.
,
Brennan
M.
,
Scherer
S.
,
Stinchcomb
D. T.
,
Struhl
K.
,
Davis
R. W.
(
1979
).
Sterile host yeasts (SHY): a eukaryotic system of biological containment for recombinant DNA experiments.
Gene
8
,
17
24
Bowser
R.
,
Muller
H.
,
Govindan
B.
,
Novick
P.
(
1992
).
Sec8p and Sec15p are components of a plasma membrane-associated 19.5s particle that may function downstream of Sec4p to control exocytosis.
J. Cell Biol
118
,
1041
1056
Brennwald
P.
,
Kearns
B.
,
Champion
K.
,
Keränen
S.
,
Bankaitis
V.
,
Novick
P.
(
1994
).
Sec9 is a SNAP-25-like component of a yeast SNARE complex that may be the effector of Sec4 function in exocytosis.
Cell
79
,
245
258
Bretscher
M. S.
,
Munro
S.
(
1993
).
Cholesterol and the Golgi apparatus.
Science
261
,
1280
1281
Cereghino
J. L.
,
Marcusson
E. G.
,
Emr
S. D.
(
1995
).
The cytoplasmic tail domain of the vacuolar protein sorting receptor Vps10p and a subset of VPS gene products regulate receptor stability, function, and localization.
Mol. Biol. Cell
6
,
1089
1102
Cooper
A. A.
,
Stevens
T. H.
(
1996
).
Vps10p cycles between the late-Golgi and prevacuolar compartments in its function as the sorting receptor for multiple yeast vacuolar hydrolases.
J. Cell Biol
133
,
529
541
Dascher
C.
,
Ossig
R.
,
Gallwitz
D.
,
Schmitt
H. D.
(
1991
).
Identification and structure of four yeast genes (SLY) that are able to suppress the functional loss of YPT1, a member of the RAS superfamily.
Mol. Cell. Biol
11
,
872
885
Franzusoff
A.
,
Schekman
R.
(
1989
).
Functional compartments of the yeast Golgi apparatus are defined by the sec7 mutation.
EMBO J
8
,
2695
2702
Franzusoff
A.
,
Lauze
E.
,
Howell
K. E.
(
1992
).
Immuno-isolation of Sec7p-coated transport vesicles from the yeast secretory pathway.
Nature
355
,
173
175
Goud
B.
,
Zahraoui
A.
,
Tavitian
A.
,
Saraste
J.
(
1990
).
Small GTP-binding protein associated with Golgi cisternae.
Nature
345
,
553
556
Hengst
L.
,
Lehmeier
T.
,
Gallwitz
D.
(
1990
).
The ryh1 gene in the fission yeast Schizosaccharomyces pombe encoding a GTP-binding protein related to ras, rho and ypt: structure, expression and identification of its human homologue.
EMBO J
9
,
1949
1955
Horazdovsky
B. F.
,
Busch
G. R.
,
Emr
S. D.
(
1994
).
VPS21 encodes a rab5-like GTP binding protein that is required for the sorting of yeast vacuolar proteins.
EMBO J
13
,
1297
1309
Ito
H.
,
Fukuda
Y.
,
Murata
K.
,
Kimura
A.
(
1983
).
Transformation of intact yeast cells treated with alkali cations.
J. Bacteriol
153
,
163
168
Jones
D. T.
,
Taylor
W. R.
,
Thornton
J. M.
(
1994
).
A model recognitionapproach to the prediction of all-helical membrane protein structure and topology.
Biochemistry
33
,
3038
3049
Julius
D.
,
Blair
L.
,
Brake
A.
,
Sprague
G.
,
Thorner
J.
(
1983
).
Yeastfactor is processed from a larger precursor polypeptide: the essential role of a membrane-bound dipeptidyl aminopeptidase.
Cell
32
,
839
852
Kaiser
C. A.
,
Schekman
R.
(
1990
).
Distinct sets of SEC genes govern transport vesicle formation and fusion early in the secretory pathway.
Cell
61
,
723
733
Klionsky
D. J.
,
Herman
P. K.
,
Emr
S. D.
(
1990
).
The fungal vacuole: composition, function and biogenesis.
Microbiol. Rev.
54
,
266
292
Koonin
E. V.
,
Bork
P.
,
Sander
C.
(
1994
).
Yeast chromosome III: new gene functions.
EMBO J
13
,
493
503
Kyte
J.
,
Doolittle
R. F.
(
1982
).
A simple method for displaying the hydropathic character of a protein.
J. Mol. Biol
157
,
105
132
Martinez
O.
,
Schmidt
A.
,
Salamero
J.
,
Hoflack
B.
,
Roa
M.
,
Goud
B.
(
1994
).
The small GTP-binding protein rab6 functions in intra-Golgi transport.
J. Cell Biol
6
,
1575
1588
Nothwehr
S. F.
,
Conibear
E.
,
Stevens
T. H.
(
1995
).
Golgi and vacuolar membrane proteins reach the vacuole in vps1 mutant yeast cells via the plasma membrane.
J. Cell Biol
129
,
35
46
Novick
P.
,
Ferro
S.
,
Schekman
R.
(
1981
).
Order of events in the yeast secretory pathway.
Cell
25
,
461
469
Oliver
S.G.
,
van der Aart
Q. J. M.
,
Agostini-Carbone
M.L.
,
Aigle
M.
,
Alberghina
M.
,
Alexandraki
D.
,
Antoine
G.
,
Anwar
R.
,
Ballesta
J.P. G.
,
Benit
P.
, et al. 
(
1992
).
The complete DNA sequence of yeast chromosome III.
Nature
357
,
38
46
Ossig
R.
,
Dascher
C.
,
Trepte
H.-H.
,
Schmitt
H. D.
,
Gallwitz
D.
(
1991
).
The yeast SLY gene products, suppressors of defects in the essential GTP-binding YPT1 protein, may act in endoplasmic reticulum-to-Golgi transport.
Mol. Cell. Biol
11
,
2980
2993
Oyler
G. A.
,
Higgins
G. A.
,
Hart
R. A.
,
Battenberg
E.
,
Billingsley
M.
,
Bloom
F. E.
,
Wilson
M. C.
(
1989
).
The identification of a novel synaptosomal-associated protein, SNAP-25, differentially expressed by neuronal subpopulations.
J. Cell Biol
109
,
3039
3052
Payne
G. S.
,
Schekman
R.
(
1989
).
Clathrin: a role in the intracellular retention of a Golgi membrane protein.
Science
245
,
1358
1365
Raymond
C. K.
,
Roberts
C. J.
,
Moore
K. E.
,
Howald
I.
,
Stevens
T. H.
(
1992
).
Biogenesis of the vacuole in Saccharomyces cerevisiae.
Int. Rev. Cytol
139
,
59
120
Raymond
C. K.
,
Howald-Stevenson
I.
,
Vater
C. A.
,
Stevens
T. H.
(
1992
).
Morphological classification of the yeast vacuolar protein sorting mutants: evidence for a prevacuolar compartment in class E vps mutants.
Mol. Biol. Cell
3
,
1389
1402
Robinson
J. S.
,
Klionsky
D. J.
,
Banta
L. M.
,
Emr
S. D.
(
1988
).
Protein sorting in Saccharomyces cerevisiae: isolation of mutants defective in the delivery and processing of multiple vacuolar hydrolases.
Mol. Cell. Biol
8
,
4936
4948
Rothman
J. E.
(
1994
).
Mechanisms of intracellular protein transport.
Nature
372
,
55
63
Salminen
A.
,
Novick
P. J.
(
1987
).
A ras -like protein is required for a post-Golgi event in yeast secretion.
Cell
49
,
527
538
Schmitt
H. D.
,
Puzicha
M.
,
Gallwitz
D.
(
1988
).
Study of a temperature-sensitive mutant of the ras-related YPT1 gene product in yeast suggests a role in the regulation of intracellular calcium.
Cell
53
,
635
647
Seeger
M.
,
Payne
G. S.
(
1992
).
Selective and immediate effects of clathrin heavy chain mutations on Golgi membrane protein retention in Saccharomyces cerevisiae.
J. Cell Biol
118
,
531
540
Segev
N.
,
Mulholland
J.
,
Botstein
D.
(
1988
).
The yeast GTP-binding YPT1 protein and a mammalian counterpart are associated with the secretion machinery.
Cell
52
,
915
924
Sikorski
R. S.
,
Hieter
P.
(
1989
).
A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae.
Genetics
122
,
19
27
Singer-Kruger
B.
,
Stenmark
H.
,
Dusterhöft
A.
,
Philippsen
P.
,
Yoo
J.-S.
,
Gallwitz
D.
,
Zerial
M.
(
1994
).
Role of three Rab5-like GTPases, Ypt51p, Ypt52p, and Ypt53p, in the endocytic and vacuolar protein sorting pathways of yeast.
J. Cell Biol
125
,
283
298
Søgaard
M.
,
Tani
K.
,
Ye
R. R.
,
Geromanos
S.
,
Tempst
P.
,
Kirchhausen
T.
,
Rothman
J. E.
,
Söllner
T.
(
1994
).
A Rab protein is required for the assembly of SNARE complexes in the docking of transport vesicles.
Cell
78
,
937
948
Söllner
T.
(
1995
).
SNAREs and targeted membrane fusion.
FEBSLett
369
,
80
83
Stack
J. H.
,
Horazdovsky
B.
,
Emr
S. D.
(
1995
).
Receptor-mediated protein sorting to the vacuole in yeast: roles for a protein kinase, a lipid kinase and GTP-binding proteins.
Annu. Rev. Cell. Dev. Biol
11
,
1
33
Stenius
K.
,
Janz
R.
,
Sudhof
T. C.
,
Jahn
R.
(
1995
).
Structure of Synaptogyrin (p29) defines novel synaptic vesicle protein.
J. Cell Biol
131
,
1801
1809
Wichmann
H.
,
Hengst
L.
,
Gallwitz
D.
(
1992
).
Endocytosis in yeast: evidence for the involvement of a small GTP-binding protein (Ypt7p).
Cell
71
,
1131
1142
Wilsbach
K.
,
Payne
G. S.
(
1993
).
Vps1p, a member of the dynamin GTPase family, is necessary for Golgi membrane protein retention in Saccharomycescerevisiae.
EMBO J
12
,
3049
3059
Zerial
M.
,
Stenmark
H.
(
1993
).
Rab GTPases in vesicular transport.
Curr. Opin. Cell Biol
5
,
613
620
This content is only available via PDF.