Specificity of Cdk activation in vivo by the two Caks Mcs6 and Csk1 in fission yeast

Hermand, Damien; Westerling, Thomas; Pihlak, Arno; Thuret, Jean‐Yves; Vallenius, Tea; Tiainen, Marianne; Vandenhaute, Jean; Cottarel, Guillaume; Mann, Carl; Mäkelä, Tomi P.

doi:10.1093/emboj/20.1.82

Cited by 32 publications

(42 citation statements)

References 69 publications

(138 reference statements)

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“…This suggests that the Pmh1 immunoprecipitation could contain other fission yeast TFIIH subunits [38]. These observations together with the demonstration that T-loop phosphorylation of Mcs6 is not essential [6] are consistent with a model where association of Pmh1 with the Mcs6 CDK and Mcs2 cyclin alleviates the requirement of T-loop phosphorylation as observed in other species [8,9,21,42], and that Pmh1 represents the Mat1 homolog in fission yeast.…”

Section: Pmh1 Is the Mat1 Homolog Of Fission Yeast And Activates The supporting

confidence: 86%

“…The resulting plasmid was introduced into Sf9 insect cells together with BaculoGold (Pharmingen) baculovirus DNA as recommended by the manufacturer. Mcs6 and Mcs2 baculoviruses have been described elsewhere [6].…”

Section: Methodsmentioning

confidence: 99%

“…Soluble protein extracts were prepared from log phase cultures as described [6]. For immunoprecipitations of FLAG-pmh1 2 ll of anti-FLAG (M5 monoclonal antibody, Eastman Kodak) and 10 ll (bead volume) of protein A-Sepharose were added to 200 lg of total protein.…”

Section: Methodsmentioning

confidence: 99%

“…Indeed, the Mcs6-Mcs2 complex is able to directly phosphorylate the T-loop activation site on Cdc2 [5]. Mcs6 also activates Cdc2 in vivo [6,7], and thus represents a CDK-activating kinase (CAK) in fission yeast.…”

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confidence: 99%

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Mcs2 and a novel CAK subunit Pmh1 associate with Skp1 in fission yeast

Bamps

Westerling

Pihlak

et al. 2004

Biochemical and Biophysical Research Communications

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The Mcs6 CDK together with its cognate cyclin Mcs2 represents the CDK-activating kinase (CAK) of fission yeast Cdc2. We have attempted to determine complexes in which Mcs6 and Mcs2 mediate this and possible other functions. Here we characterize a novel interaction between Mcs2 and Skp1, a component of the SCF (Skp1-Cullin-F box protein) ubiquitin ligase. Furthermore, we identify a novel protein termed Pmh1 through its association with Skp1. Pmh1 associates with the Mcs6-Mcs2 complex, enhancing its kinase activity, and represents the apparent homolog of metazoan Mat1. Association of Mcs2 or Pmh1 with Skp1 does not appear to be involved in proteolytic degradation, as these complexes do not contain Pcu1, and levels of Mcs2 or Pmh1 are not sensitive to inhibition of SCF and the 26S proteasome. The identified interactions between Skp1 and two regulatory CAK subunits may reflect a novel mechanism to modulate activity and specificity of the Mcs6 kinase.

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Section: Pmh1 Is the Mat1 Homolog Of Fission Yeast And Activates The supporting

confidence: 86%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Mcs2 and a novel CAK subunit Pmh1 associate with Skp1 in fission yeast

Bamps

Westerling

Pihlak

et al. 2004

Biochemical and Biophysical Research Communications

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“…These results point to a role for Csk1 as a CAK-activating kinase (34). csk1 can rescue a cak1 mutant in S. cerevisiae (35), and both Csk1 and Mcs6/Mcs2 may activate the S. pombe cdk Cdc2 in vitro, opening the possibility of redundant CAK functions (33). The pathogenic yeast Candida albicans also has a protein that resembles Cak1p.…”

mentioning

confidence: 86%

Comparison of Cak1p-like Cyclin-dependent Kinase-activating Kinases

Tsakraklides

Solomon

2002

Journal of Biological Chemistry

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Cyclin-dependent kinases (cdks) coordinate progression through the eukaryotic cell cycle and require phosphorylation by a cdk-activating kinase (CAK) for full activity. In most eukaryotes Cdk7 is the catalytic subunit of a heterotrimeric CAK (Cdk7-cyclin H-Mat1) that is also involved in transcription as part of the transcription factor IIH complex. The Saccharomyces cerevisiae CAK, Cak1p, is a monomeric protein kinase with an atypical sequence and unusual biochemical properties compared with trimeric CAKs and other protein kinases. We sought to determine whether these properties were shared by a small group of monomeric CAKs that can function in place of CAK1 in S. cerevisiae. We found that Schizosaccharomyces pombe Csk1, Candida albicans Cak1, and Arabidopsis thaliana Cak1At, like Cak1p, all displayed a preference for cyclin-free cdk substrates, were insensitive to the protein kinase inhibitor 5 -fluorosulfonylbenzoyladenosine (FSBA), and were insensitive to mutation of a highly conserved lysine residue found in the nucleotide binding pocket of all protein kinases. The S. pombe and C. albicans kinases also resembled Cak1p in their kinetics of nucleotide and protein substrate utilization. Conservation of these unusual properties in fungi and plants points to shared evolutionary requirements not met by Cdk7 and raises the possibility of developing antifungal agents targeting CAKs.

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