Regulation of the Cln3-Cdc28 kinase by cAMP in Saccharomyces cerevisiae

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.

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An Altered Nuclear Migration into the Daughter Bud is Induced by the Cyclin A1-mediated Cdc28 Kinase through an Aberrant Spindle Movement in Saccharomyces cerevisiae.

Cell Structure and Function ◽

10.1247/csf.22.465 ◽

1997 ◽

Vol 22 (4) ◽

pp. 465-476 ◽

Cited By ~ 3

Author(s):

Hashmat Sikder ◽

Minoru Funakoshi ◽

Takeharu Nishimoto ◽

Hideki Kobayashi

Keyword(s):

Saccharomyces Cerevisiae ◽

Nuclear Migration ◽

Cyclin A1 ◽

Cdc28 Kinase

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Expression of p60v-src in Saccharomyces cerevisiae results in elevation of p34CDC28 kinase activity and release of the dependence of DNA replication on mitosis.

Molecular and Cellular Biology ◽

10.1128/mcb.13.8.5112 ◽

1993 ◽

Vol 13 (8) ◽

pp. 5112-5121 ◽

Cited By ~ 10

Author(s):

F Boschelli

Keyword(s):

Saccharomyces Cerevisiae ◽

Dna Replication ◽

Dna Content ◽

Protein Tyrosine Kinase ◽

Flow Cytometric Analysis ◽

Yeast Saccharomyces Cerevisiae ◽

Genetic Studies ◽

Flow Cytometric ◽

Oncogenic Protein ◽

Cdc28 Kinase

Expression of the oncogenic protein tyrosine kinase p60v-src in the yeast Saccharomyces cerevisiae has been shown to result in rapid cell death (J. S. Brugge, G. Jarosik, J. Andersen, A. Queral-Lustig, M. Fedor-Chaiken, and J. R. Broach, Mol. Cell. Biol. 7:2180-2187, 1987). Work described here demonstrates that v-Src expression results in accumulation of large-budded cells and a nuclear division block without blocking cytokinesis. Flow-cytometric analysis indicates that the DNA content of these cells is elevated beyond the G2 DNA content, and genetic studies indicate that v-Src expression causes aneuploidy. The activity of Cdc28 kinase, which controls the G1/S and G2/M transitions in S. cerevisiae, increases during galactose induction in a Src+ strain but not in an isogenic Src- strain. These observations indicate that v-Src expression disrupts p34CDC28 kinase regulation, allowing DNA replication to proceed in the absence of a prior mitotic event.

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Three independent forms of regulation affect expression of HO, CLN1 and CLN2 during the cell cycle of Saccharomyces cerevisiae.

Genetics ◽

10.1093/genetics/138.4.1015 ◽

1994 ◽

Vol 138 (4) ◽

pp. 1015-1024 ◽

Cited By ~ 5

Author(s):

L Breeden ◽

G Mikesell

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Cycle ◽

Normal Cell ◽

Cell Cycle Regulation ◽

Cell Cycle Control ◽

G1 Cyclins ◽

Affect Expression ◽

Cdc28 Kinase ◽

Cycle Regulation ◽

Normal Cell Cycle

Abstract The G1 cyclins (CLNs) bind to and activate the CDC28 kinase during the G1 to S transition in Saccharomyces cerevisiae. Two G1 cyclins are regulated at the RNA level so that their RNAs peak at the G1/S boundary. In this report we show that the cell cycle regulation of CLN1 and CLN2 is partially determined by the restricted expression of SW14, a known trans-activator of SCB elements. When SWI4 is constitutively expressed or deleted, cell cycle regulation of CLN1/2 is reduced but not eliminated. In the absence of SwI6, another known regulator of both SCB and MCB elements, cell cycle regulation of the CLNs is also reduced, and the Start-dependence of HO transcription is eliminated. This indicates that SwI6 also plays an important role in the normal cell cycle regulation of all three promoters. When both SwI6 activity and the transcriptional regulation of SW14 are eliminated, cell cycle regulation is further reduced, indicating that these are two independent pathways of regulation. However, a twofold fluctuation in transcript levels still persists under these conditions. This reveals a third source of cell cycle control, which could affect SwI4 activity post-transcriptionally, or reflect the existence of another unidentified regulator of these promoters.

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p34Cdc28-mediated control of Cln3 cyclin degradation.

Molecular and Cellular Biology ◽

10.1128/mcb.15.2.731 ◽

1995 ◽

Vol 15 (2) ◽

pp. 731-741 ◽

Cited By ~ 193

Author(s):

J Yaglom ◽

M H Linskens ◽

S Sadis ◽

D M Rubin ◽

B Futcher ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

G1 Phase ◽

Phosphorylation Sites ◽

Nonpermissive Temperature ◽

Yeast Saccharomyces Cerevisiae ◽

Rapid Degradation ◽

Ubiquitin Conjugating Enzyme ◽

A Cell ◽

Cdc28 Kinase ◽

Beta Galactosidase

Cln3 cyclin of the budding yeast Saccharomyces cerevisiae is a key regulator of Start, a cell cycle event in G1 phase at which cells become committed to division. The time of Start is sensitive to Cln3 levels, which in turn depend on the balance between synthesis and rapid degradation. Here we report that the breakdown of Cln3 is ubiquitin dependent and involves the ubiquitin-conjugating enzyme Cdc34 (Ubc3). The C-terminal tail of Cln3 functions as a transferable signal for degradation. Sequences important for Cln3 degradation are spread throughout the tail and consist largely of PEST elements, which have been previously suggested to target certain proteins for rapid turnover. The Cln3 tail also appears to contain multiple phosphorylation sites, and both phosphorylation and degradation of Cln3 are deficient in a cdc28ts mutant at the nonpermissive temperature. A point mutation at Ser-468, which lies within a Cdc28 kinase consensus site, causes approximately fivefold stabilization of a Cln3-beta-galactosidase fusion protein that contains a portion of the Cln3 tail and strongly reduces the phosphorylation of this protein. These data indicate that the degradation of Cln3 involves CDC28-dependent phosphorylation events.

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Phosphorylation of Sic1, a Cyclin-dependent Kinase (Cdk) Inhibitor, by Cdk Including Pho85 Kinase Is Required for Its Prompt Degradation

Molecular Biology of the Cell ◽

10.1091/mbc.9.9.2393 ◽

1998 ◽

Vol 9 (9) ◽

pp. 2393-2405 ◽

Cited By ~ 70

Author(s):

Masafumi Nishizawa ◽

Masaoki Kawasumi ◽

Marie Fujino ◽

Akio Toh-e

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Growth ◽

Cdk Inhibitor ◽

Phosphorylation Sites ◽

Cyclin Dependent Kinase ◽

Yeast Saccharomyces Cerevisiae ◽

Cdk Phosphorylation ◽

Cdc28 Kinase

In the yeast Saccharomyces cerevisiae, Sic1, an inhibitor of Clb-Cdc28 kinases, must be phosphorylated and degraded in G1for cells to initiate DNA replication, and Cln-Cdc28 kinase appears to be primarily responsible for phosphorylation of Sic1. The Pho85 kinase is a yeast cyclin-dependent kinase (Cdk), which is not essential for cell growth unless both CLN1 andCLN2 are absent. We demonstrate that Pho85, when complexed with Pcl1, a G1cyclin homologue, can phosphorylate Sic1 in vitro, and that Sic1 appears to be more stable inpho85Δ cells. Three consensus Cdk phosphorylation sites present in Sic1 are phosphorylated in vivo, and two of them are required for prompt degradation of the inhibitor. Pho85 and other G1Cdks appear to phosphorylate Sic1 at different sites in vivo. Thus at least two distinct Cdks can participate in phosphorylation of Sic1 and may therefore regulate progression through G1.

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Expression of p60v-src in Saccharomyces cerevisiae results in elevation of p34CDC28 kinase activity and release of the dependence of DNA replication on mitosis

Molecular and Cellular Biology ◽

10.1128/mcb.13.8.5112-5121.1993 ◽

1993 ◽

Vol 13 (8) ◽

pp. 5112-5121

Author(s):

F Boschelli

Keyword(s):

Saccharomyces Cerevisiae ◽

Dna Replication ◽

Dna Content ◽

Protein Tyrosine Kinase ◽

Flow Cytometric Analysis ◽

Yeast Saccharomyces Cerevisiae ◽

Genetic Studies ◽

Flow Cytometric ◽

Oncogenic Protein ◽

Cdc28 Kinase

Expression of the oncogenic protein tyrosine kinase p60v-src in the yeast Saccharomyces cerevisiae has been shown to result in rapid cell death (J. S. Brugge, G. Jarosik, J. Andersen, A. Queral-Lustig, M. Fedor-Chaiken, and J. R. Broach, Mol. Cell. Biol. 7:2180-2187, 1987). Work described here demonstrates that v-Src expression results in accumulation of large-budded cells and a nuclear division block without blocking cytokinesis. Flow-cytometric analysis indicates that the DNA content of these cells is elevated beyond the G2 DNA content, and genetic studies indicate that v-Src expression causes aneuploidy. The activity of Cdc28 kinase, which controls the G1/S and G2/M transitions in S. cerevisiae, increases during galactose induction in a Src+ strain but not in an isogenic Src- strain. These observations indicate that v-Src expression disrupts p34CDC28 kinase regulation, allowing DNA replication to proceed in the absence of a prior mitotic event.

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