The cdc2-related protein p40MO15 is the catalytic subunit of a protein kinase that can activate p33cdk2 and p34cdc2.

Abstract

Activation of the cyclin‐dependent protein kinases p34cdc2 and p33cdk2 requires binding with a cyclin partner and phosphorylation on the first threonine residue in the sequence THEVVTLWYRAPE. We present evidence that this threonine residue, number 160 in p33cdk2, can be specifically phosphorylated by a cdc2‐related protein kinase from Xenopus oocytes called p40MO15. Binding to cyclin A and phosphorylation of this threonine are both required to activate fully the histone H1 kinase activity of p33cdk2. In cell extracts, a portion of p40MO15 is found in a high molecular weight complex that is considerably more active than a lower molecular weight form. Wild‐type MO15 protein expressed in bacteria does not possess kinase activity, but acquires p33cdk2‐T160 kinase activity after incubation with cell extract and ATP. We conclude that p40MO15 corresponds to CAK (cdc2/cdk2 activating kinase) and speculate that, like p33cdk2 and p34cdc2, p40MO15 requires activation by phosphorylation and association with a companion subunit.

Interferon-induces expression of cyclin-dependent kinase-inhibitors p21WAF1 and p27Kip1 that prevent activation of cyclin-dependent kinase by CDK-activating kinase (CAK)

“…The observed IFN-a-mediated early inhibition (6 ± 24 h treatment) of CDK-4-associated H-1 kinase activity was not due to the inhibitory e ect of CDK-4 expression, as IFN-atreatment did not inhibit the expression of CDK-4 protein upto 24 h treatment ( Figure 2c). Results in Figure 2d show that the induced p21 WAF1 also interacts with CDK-4, in addition to the reported interaction with CDK-2 (Poon et al, 1993;Matsuoka et al, 1994). Since the kinetics of restoration (due to depletion of p21 WAF1 ) of H-1 activity associated with CDK-2 or CDK-4 was in-parallel with the kinetics of p21 WAF1 induction, these results suggested that the observed inhibition of CDK-2 or CDK-4 kinase activity in IFNa-treated Daudi cells may be due to conditional induction of CDK-inhibitor p21 WAF1 by IFN-a.…”

“…One of the key phosphorylation event controlling CDKs activity is the activation of phosphorylation on a residue in various CDKs, corresponding to threonine (Thr) 161 in human prototypic CDK-1. Phosphorylation of this residue is positively required for function of both human and yeast CDK-1 and recent reports have shown that an analogous phosphorylation step is essential for activities of CDK-2 and CDK-4 (Poon et al, 1993;. A kinase responsible for phosphorylating the Thr 161 of CDK-1 and the corresponding residue in other CDKs (T 172 in CDK-4, T 160 in CDK-2) has been identi®ed in various organisms and designated CAK, for CDK-activating kinase (also known as CDK-7).…”

Expression of a novel form of p21Cip1/Waf1 in UV-irradiated and transformed cells

“…GST-Cdk2 (human) in pGEX-2T was as described (Poon et al, 1993), and cdk2 (Xenopus) in pET8c for reticulocyte lysate translation was a gift from T Hunt (ICRF, South Mimms). GST-tagged human p21 (GST-p21 in pGEX-KG) and histidine-tagged human p21 (p21-H6 in pET21d) were as described (Poon et al, 1996).…”

“…Thrombin digestion of GST-fusion proteins was as described (Poon et al, 1993). Transcription of mRNA in vitro and translation in the mRNA-dependent reticulocyte lysate in the presence of 35 Smethionine was as described (Jackson and Hunt, 1983).…”

Quiescence versus apoptosis: Myc abundance determines pathway of exit from the cell cycle

“…A C-terminal fragment of the human retinoblastoma (Rb) protein (aa 773 ± 928) fused to glutathione-S-transferase (GST), encoded by the pGEX-GST-C-Rb plasmid (provided by E Harlow and W Kaelin) was isolated as described (Poon et al, 1993). Tet-myc cells were trypsinized, pelleted and quick frozen in liquid nitrogen at various times following serum starvation.…”