The lethal effects of the expression of the oncogenic protein tyrosine kinase p60v-src in Saccharomyces cerevisiae are associated with a loss of cell cycle control at the G1/S and G2/M checkpoints. Results described here indicate that the ability of v-Src to kill yeast is dependent on the integrity of the SH2 domain, a region of the Src protein involved in recognition of proteins phosphorylated on tyrosine. Catalytically active v-Src proteins with deletions in the SH2 domain have little effect on yeast growth, unlike wild-type v-Src protein, which causes accumulation of large-budded cells, perturbation of spindle microtubules and increased DNA content when expressed. The proteins phosphorylated on tyrosine in cells expressing v-Src differ from those in cells expressing a Src protein with a deletion in the SH2 domain. Also, unlike the wild-type v-Src protein, which drastically increases histone H1-associated Cdc28 kinase activity, c-Src and an altered v-Src protein have no effect on Cdc28 kinase activity. These results indicate that the SH2 domain is functionally important in the disruption of the yeast cell cycle by v-Src.

Adams
A. E. M.
,
Pringle
J. R.
(
1984
).
Relationship of actin and tubulin distribution to bud growth in wild-type and morphogenetic-mutant Saccharomyces cerevisiae.
J. Cell Biol
98
,
934
945
Bagrodia
S.
,
Chackalaparampil
I.
,
Kmiecik
T. E.
,
Shalloway
D.
(
1991
).
Altered tyrosine 527 phosphorylation and mitotic activation of p60c-src.
Nature
349
,
172
175
Brizuela
L.
,
Draetta
G.
,
Beach
D.
(
1987
).
p13suc1 acts in the fission yeast cell division cycle as a component of the p34cdc2 protein kinase.
EMBO J
6
,
3507
3514
Brugge
J. S.
,
Jarosik
G.
,
Andersen
J.
,
Queral-Lustig
A.
,
Fedor-Chaiken
M.
,
Broach
J. R.
(
1987
).
Expression of Rous sarcoma virus transforming protein pp60v-src in Saccharomyces cerevisiae cells.
Mol. Cell. Biol
7
,
2180
2187
Bryant
D. L.
,
Parsons
J. T.
(
1984
).
Amino acid alterations within a highly conserved region of the Rous sarcoma virus src gene product pp60src inactivate tyrosine protein kinase activity.
Mol. Cell. Biol
4
,
862
866
Chackalaparampil
I.
,
Shalloway
D.
(
1988
).
Altered phosphorylation and activation of pp60c-src during fibroblast mitosis.
Cell
52
,
801
810
Cooper
J. A.
,
Gould
K. L.
,
Cartwright
C. A.
,
Hunter
T.
(
1986
).
Tyr527 is phosphorylated in pp60c-src: implications for regulation.
Science
231
,
1431
1434
Cooper
J. A.
,
King
C. S.
(
1986
).
Dephosphorylation or antibody528F. Boschelli, S. M. Uptain and J. J. Lightbodybinding to the carboxy terminus stimulates pp60 c-src.
Mol. Cell. Biol
6
,
4467
4477
Cooper
J. A.
,
Runge
K.
(
1987
).
Avian pp60c-src is more active when expressed in yeast than in vertebrate fibroblasts.
Oncogene Res
1
,
297
–.
Courtneidge
S. A.
(
1985
).
Activation of the pp60c-src kinase by middle T antigen binding or by dephosphorylation.
EMBO J
4
,
1471
1477
Cross
F. R.
,
Garber
E. A.
,
Hanafusa
H.
(
1985
).
N-terminal deletions in Rous sarcoma virus p60v-src: effects on tyrosine kinase and biological activities and on recombination in tissue culture with the cellular src gene.
Mol. Cell. Biol
5
,
2789
2795
Forsburg
S. L.
,
Nurse
P.
(
1991
).
Cell cycle regulation in the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe.
Annu. Rev. Cell Biol
7
,
227
256
Fukui
Y.
,
O'Brien
M. C.
,
Hanafusa
H.
(
1991
).
Deletions in the SH2 domain of p60v-src prevent association with the detergent-insoluble cellular matrix.
Mol. Cell. Biol
11
,
1207
1213
Hamaguchi
M.
,
Hanafusa
H.
(
1987
).
Association of p60src with Triton X-100 resistant cellular structure correlates with morphological transformation.
Proc. Nat. Acad. Sci. USA
84
,
2312
2316
Hirai
H.
,
Varmus
H. E.
(
1990
).
Site-directed mutagenesis of the SH2-and SH3-coding domains of c-src produces varied phenotypes, including oncogenic activation of p60v-src.
Mol. Cell. Biol
10
,
1307
1318
Hoyt
M. A.
,
Totis
L.
,
Roberts
B. T.
(
1991
).
S. cerevisiae genes required for cell cycle arrest in response to loss of microtubule function.
Cell
66
,
507
517
Itoh
H.
,
Fukuda
Y.
,
Murata
K.
,
Kimura
A.
(
1983
).
Transformation of intact yeast cells treated with alkali cations.
J. Bacteriol
153
,
163
168
Kanik-Ennulat
C.
,
Neff
N.
(
1990
).
Vanadate-resistant mutants of Saccharomyces cerevisiae show alterations in protein phosphorylation and growth control.
Mol. Cell. Biol
10
,
898
909
Kanner
S. B.
,
Reynolds
A. B.
,
Wang
H.-C. R.
,
Vines
R. R.
,
Parsons
J. T.
(
1991
).
The SH2 and SH3 domains of pp60src direct stable association with tyrosine phosphorylated proteins p130 and p110.
EMBO J
10
,
1689
1698
Kaplan
J. M.
,
Varmus
H. E.
,
Bishop
J. M.
(
1990
).
The src protein contains multiple domains for specific attachment to membranes.
Mol. Cell. Biol
10
,
1000
1009
Koch
C. A.
,
Anderson
D.
,
Moran
M. F.
,
Ellis
C.
,
Pawson
T.
(
1991
).
SH2 and SH3 domains: Elements that control interactions of cytoplasmic signalling proteins.
Science
252
,
668
674
Li
R.
,
Murray
A. W.
(
1991
).
Feedback control of mitosis in budding yeast.
Cell
66
,
519
531
Lipsich
L. A.
,
Lewis
A. J.
,
Brugge
J. L.
(
1983
).
Isolation of monoclonal antibodies that recognize the transforming proteins of avian sarcoma viruses.
J. Virol
48
,
352
360
Matsuda
M.
,
Mayer
B. J.
,
Hanafusa
H.
(
1991
).
Association ofp47gag-crk with phosphotyrosine-containing proteins and tyrosine kinase activity through the SH2 domain.
Mol. Cell. Biol
11
,
1607
1613
Mayer
B. J.
,
Jackson
P. K.
,
Baltimore
D.
(
1991
).
The noncatalytic src homolgy region 2 segment of abl tyrosine kinase binds to tyrosine-phosphorylated cellular proteins with high affinity.
Proc. Nat. Acad. Sci. USA
88
,
627
631
Mayer
B. J.
,
Jackson
P. K.
,
Elten
R. A. V.
,
Baltimore
D.
(
1992
).
Point mutations in the abl SH2 domain coordinately impair phosphotyrosine binding in vitro and transforming activity in vivo.
Mol. Cell. Biol
12
,
609
618
Nemeth
S. P.
,
Fox
L. G.
,
DeMarco
M.
,
Brugge
J. S.
(
1989
).
Deletions within the amino-terminal half of the c-src gene product that alter the functional activity of the protein.
Mol. Cell. Biol
9
,
1109
1119
O'Brien
M. C.
,
Fukui
Y.
,
Hanafusa
H.
(
1990
).
Activation of the proto-oncogene p60c-src by point mutations in the SH2 domain.
Mol. Cell. Biol
10
,
2855
2862
Overduin
M.
,
Rios
C. B.
,
Mayer
B. J.
,
Baltimore
D.
,
Cowburn
D.
(
1992
).
Three-dimensional solution structure of the src homology 2 domain of c-abl.
Cell
70
,
697
704
Piwnica-Worms
H.
,
Saunders
K. B.
,
Roberts
T. M.
,
Smith
A. E.
,
Cheng
S. H.
(
1987
).
Tyrosine phosphorylation regulates the biochemical and biological properties of pp60c-src.
Cell
49
,
75
82
Reid
G. A.
(
1983
).
Pulse-labeling of yeast cells and spheroplasts.
Meth. Enzymol
97
,
324
329
Roussel
R. R.
,
Brodeur
S. R.
,
Shalloway
D.
,
Laudano
A. P.
(
1991
).
Selective binding of activated pp60c-src by an immobilized synthetic phosphopeptide modeled on the carobxyl terminus of pp60c-src.
Proc. Nat. Acad. Sci. USA
88
,
10696
10700
Schwartz
D. E.
,
Tizard
R.
,
Gilbert
W.
(
1983
).
Nucleotide sequence of Rous sarcoma virus.
Cell
32
,
853
869
Waksman
G.
,
Kominos
D.
,
Robertson
S. C.
,
Pant
N.
,
Baltimore
D.
,
Birge
R. B.
,
Cowburn
D.
,
Hanafusa
H.
,
Mayer
B. J.
,
Overduin
M.
,
Resh
M. D.
,
Rios
C. B.
,
Silverman
L.
,
Kuriyan
J.
(
1992
).
Crystal structure of the phosphotyrosine recognition domain SH2 of v-src complexed with tyrosine-phosphorylated peptides.
Nature
358
,
646
653
Wang
H.-C. R.
,
Parsons
J. T.
(
1989
).
Deletions and insertions within an amino-terminal domain of pp60v-src inactivate transformation and modulate membrane stability.
J. Virol
63
,
291
302
Wendler
P. A.
,
Boschelli
F.
(
1989
).
Src homology 2 domain deletion mutants of p60v-src do not phosphorylate cellular proteins of 120–150 kDa.
Oncogene
4
,
231
236
This content is only available via PDF.