Phosphorylation at Thr167 is required for Schizosaccharomyces pombe P34cdc2 function

Gould, Kathleen L.; Moreno, Sergio; Owen, David J.; Sazer, Shelley; Nurse, P.

doi:10.1016/0962-8924(92)90158-j

Cited by 144 publications

(220 citation statements)

References 57 publications

(110 reference statements)

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“…Activation of CDK6 by KSHV-cyclin is unconventional and very strong. So far all cell cycle cdk/cyclin complexes examined have required phosphorylation of the activating threonine within the T-loop by CAK for full activity (Gould et al, 1991;Desai et al, 1992;Solomon et al, 1992; ConnellCrowley et al, 1993;Fisher and Morgan, 1994;Kato et al, 1994;Matsuoka et al, 1994;Aprelikova et al, 1995;Kaldis et al, 1996. The only exception is CDK7 and its orthologs, where binding of the assembly factor MAT1 to the CDK7/cyclin H complex can substitute for activating phosphorylation Kimmelman et al, 1999).…”

Section: Discussionmentioning

confidence: 99%

“…The only exception is CDK7 and its orthologs, where binding of the assembly factor MAT1 to the CDK7/cyclin H complex can substitute for activating phosphorylation Kimmelman et al, 1999). Studies in yeast demonstrated that mutation of the activating threonine of Cdc28p (in Saccharomyces cerevisiae) or of Cdc2 (in Schizosaccharomyces pombe) rendered yeast cells inviable (Gould et al, 1991;Cismowski et al, 1995), demonstrating that the activating phosphorylation was essential. How does activation of CDK6 by KSHV-cyclin, but not by cellular D-type cyclins, bypass this requirement?…”

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confidence: 99%

“…Phosphorylation of an activating threonine (Thr-160 in CDK2 and Thr-177 in CDK6) by the cdk-activating kinase (CAK; reviewed by Kaldis, 1999) is accompanied by structural changes in the T-loop (also called the activation segment), allowing the phosphate group to interact with several other residues and thereby acting as an organizing center in the catalytic cleft (Russo et al, 1996b). Mutation of the acti- vating threonine to an unphosphorylatable amino acid prevents activation of vertebrate cdks (Desai et al, 1992;Solomon et al, 1992;Connell-Crowley et al, 1993;Kato et al, 1994;Matsuoka et al, 1994) and equivalent mutants are unable to support growth in yeast (Gould et al, 1991;Cismowski et al, 1995). Two very different CAKs have been identified.…”

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CAK-independent Activation of CDK6 by a Viral Cyclin

Kaldis

Ojala

Tong

et al. 2001

MBoC

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In normal cells, activation of cyclin-dependent kinases (cdks) requires binding to a cyclin and phosphorylation by the cdk-activating kinase (CAK). The Kaposi's sarcoma-associated herpesvirus encodes a protein with similarity to D-type cyclins. This KSHV-cyclin activates CDK6, alters its substrate specificity, and renders CDK6 insensitive to inhibition by the cdk inhibitor p16 INK4a . Here we investigate the regulation of the CDK6/KSHV-cyclin kinase with the use of purified proteins and a cell-based assay. We find that KSHV-cyclin can activate CDK6 independent of phosphorylation by CAK in vitro. In addition, CAK phosphorylation decreased the p16 INK4a sensitivity of CDK6/KSHV-cyclin complexes. In cells, expression of CDK6 or to a lesser degree of a nonphosphorylatable CDK6 T177A together with KSHV-cyclin induced apoptosis, indicating that CDK6 activation by KSHV-cyclin can proceed in the absence of phosphorylation by CAK in vivo. Coexpression of p16 partially protected cells from cell death. p16 and KSHV-cyclin can form a ternary complex with CDK6 that can be detected by binding assays as well as by conformational changes in CDK6. The Kaposi's sarcoma-associated herpesvirus has adopted a clever strategy to render cell cycle progression independent of mitogenic signals, cdk inhibition, or phosphorylation by CAK. INTRODUCTIONThe sequential activation of cyclin-dependent kinases (cdks) promotes cell cycle transitions. CDK4 and CDK6 bind to D-type cyclins and are active in G1, CDK2/cyclin E complexes function in late G1, CDK2/cyclin A complexes function in S phase, and CDC2/cyclin B and A complexes function in G2/M. The activities of cdks are regulated by protein-protein interactions (with cyclins, inhibitors, and assembly factors), protein degradation, transcriptional control, subcellular localization, and multiple phosphorylations (Pines, 1995;Sherr and Roberts, 1995;King et al., 1996;Solomon and Kaldis, 1998). Viral infection frequently targets downstream targets of cdks, resulting in inappropriate cell cycle progression.Cyclin binding activates cdks by inducing conformational changes in the structure of cdks (Jeffrey et al., 1995;Pavletich, 1999). Cyclins are unstable proteins that are synthesized and degraded periodically during the cell cycle. Transcriptional control (Koch and Nasmyth, 1994) and ubiquitin-mediated degradation (King et al., 1996) ensure the proper and irreversible timing of cell cycle regulatory events. Cyclins have also been shown to affect the substrate specificity of cdks (Peeper et al., 1993;Kelly et al., 1998;Schulman et al., 1998;Cross et al., 1999).Maximal activation of cdks requires phosphorylation of certain residues as well as dephosphorylation of others. The dual-specificity phosphatase CDC25 removes phosphates from inhibitory phosphorylation sites (Thr-14 and Tyr-15 in human CDK2) that have been phosphorylated by the WEE1/MYT1 protein kinases . Phosphorylation of an activating threonine (Thr-160 in CDK2 and Thr-177 in CDK6) by the cdk-activating kinase (CAK; reviewed by Kaldis, 199...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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confidence: 99%

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CAK-independent Activation of CDK6 by a Viral Cyclin

Kaldis

Ojala

Tong

et al. 2001

MBoC

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“…Mutants with a substitution of the threonine residue with alanine exhibit no kinase activity in vitro (Gould et al 1991;Deshaies & Kirschner 1995), and fail to suppress the temperature-sensitive growth of cdc28 or cdc2 mutants in yeast (Deshaies & Kirschner 1995;Gould et al 1991), indicating that the threonine residue is essential for the activity of these Cdks. Phosphorylation of the threonine has been demonstrated both in vivo (Gould et al 1991) and in vitro (Desai et al 1992;Solomon et al 1992). Recently, Cak1p kinase that phosphorylates T169 of Cdc28 (Cdk-activating kinase, CAK) was isolated from yeast (Kaldis et al 1996;Thuret et al 1998).…”

Section: Regulation Of Pho85p Appears Different From Other Cdksmentioning

confidence: 99%

The Pho85 kinase, a member of the yeast cyclin‐ dependent kinase (Cdk) family, has a regulation mechanism different from Cdks functioning throughout the cell cycle

et al. 1999

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Background: The PHO85 gene is a negative regulator of the PHO system in the yeast Saccharomyces cerevisiae and encodes a protein kinase (Pho85p) which is highly homologous to the Cdc28 kinase (Cdc28p). Although the two kinases share a 51% identity and their functional domains are well conserved, PHO85 fails to replace CDC28. Pho85p forms complexes with G 1 -cyclin homologues, including Pcl1p, Pcl2p and Pcl9p, and is thought to be involved in the cell-cycle regulation at G 1 and the end of M. By analysing the genetic and biochemical properties of Pho85p, we studied whether the regulation of Pho85p activity is similar to other cyclin-dependent kinases (Cdks) directly involved in cell cycle regulation.

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“…Total cell extracts of S. pombe were prepared in NP-40 buffer (Gould et al, 1991), and immunoprecipitations were carried out using either 12CA5 (anti-HA) or 9E10 (anti-Myc) antibodies as described (McDonald et al, 1999). Gel filtration was performed on protein lysates prepared in buffer A (20 mM Tris, pH 7.5, 20% glycerol, 0.1 mM EDTA, 1 mM ␤-mercaptoethanol, 5 mM ATP plus protease inhibitor cocktail [Roche, Mannheim, Germany]) using a Superose-6 column (Amersham Pharmacia Biotech, Piscataway, NJ) by FPLC.…”

Section: Protein Methodsmentioning

confidence: 99%

Identification and Characterization of Two Novel Proteins Affecting Fission Yeast γ-tubulin Complex Function

Venkatram

Tasto

Feoktistova

et al. 2004

MBoC

114

191

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The ␥-tubulin complex, via its ability to organize microtubules, is critical for accurate chromosome segregation and cytokinesis in the fission yeast, Schizosaccharomyces pombe. To better understand its roles, we have purified the S. pombe ␥-tubulin complex. Mass spectrometric analyses of the purified complex revealed known components and identified two novel proteins (i.e., Mbo1p and Gfh1p) with homology to ␥-tubulin-associated proteins from other organisms. We show that both Mbo1p and Gfh1p localize to microtubule organizing centers. Although cells deleted for either mbo1 ؉ or gfh1 ؉ are viable, they exhibit a number of defects associated with altered microtubule function such as defects in cell polarity, nuclear positioning, spindle orientation, and cleavage site specification. In addition, mbo1⌬ and gfh1⌬ cells exhibit defects in astral microtubule formation and anchoring, suggesting that these proteins have specific roles in astral microtubule function. This study expands the known roles of ␥-tubulin complex components in organizing different types of microtubule structures in S. pombe.

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Phosphorylation at Thr167 is required for Schizosaccharomyces pombe P34cdc2 function

Cited by 144 publications

References 57 publications

CAK-independent Activation of CDK6 by a Viral Cyclin

CAK-independent Activation of CDK6 by a Viral Cyclin

The Pho85 kinase, a member of the yeast cyclin‐ dependent kinase (Cdk) family, has a regulation mechanism different from Cdks functioning throughout the cell cycle

Identification and Characterization of Two Novel Proteins Affecting Fission Yeast γ-tubulin Complex Function

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