Expression and activity of p40MO15, the catalytic subunit of cdk-activating kinase, during Xenopus oogenesis and embryogenesis.

Abstract

Threonine 161 phosphorylation of p34cdc2 and its equivalent threonine 160 in p33cdk2 by cdk-activating kinase (CAK) is essential for the activation of these cyclin-dependent kinases. We have studied the expression and associated kinase activity of p40MOl5, the catalytic subunit of CAK, during Xenopus oogenesis, meiotic maturation, and early development to understand in more detail how cdk kinases are regulated during these events. We find that p40M°15 is a stable protein with a half-life >16 h that is accumulated during oogenesis. p4OMOl5 protein and its associated CAK activity are localized predominantly to the germinal vesicle; however, a small but significant proportion is found in the cytoplasm. The amount of p4oMO15 detected in stage VI oocytes remains unchanged through meiotic maturation, fertilization, and early embryogenesis. Significantly, p4OMO15 was found to be constitutively active during oogenesis, meiotic maturation, and the rapid mitotic cycles of early development. This suggests that regulation of p34cdc2 and p33cdk2 activity during cell cycle progression does not involve changes in the level or activity of p4OMOl5/CAK.

Expression of CDK7/CAK in normal and tumour cells of diverse histogenesis, cell-cycle position and differentiation

“…This question seems even more appropriate given the reports that the protein partners which bind to CDK7-namely, the cyclin H and the p36 RING finger assembly factor-also do not appreciably oscillate during the cell cycle (Devault et al, 1995;Fisher et al, 1995;Tassan et al, 1994Tassan et al, , 1995. One clue is provided by the accumulating evidence for an apparently universal role of CDK7/CAK in performing the stimulatory threonine phosphorylation of distinct CDKs operating during GI, S and G2/M phases of the somatic cell cycle (Clarke, 1995;Matsuoka et al, 1994;Morgan, 1995). Such a broad function is consistent with the ubiquitous expression in all cell types (this study), nuclear location (Darbon et al, 1994;Tassan et al, 1994; this study) and constant levels throughout the cycle (Matsuoka et al, 1994;Poon et al, 1994;Tassan et al, 1994;this study).…”

“…Such a broad function is consistent with the ubiquitous expression in all cell types (this study), nuclear location (Darbon et al, 1994;Tassan et al, 1994; this study) and constant levels throughout the cycle (Matsuoka et al, 1994;Poon et al, 1994;Tassan et al, 1994;this study). Although the activity of CAK itself is unlikely to be rate-limiting for cell-cycle progression (Matsuoka et al, 1994;Poon et aL, 1994;Tassan et al, 1994), limited access to its substrates, due to either subcellular compartmentalisation or interference by various CDK inhibitors whose function may be exerted through preventing the activatory phosphorylation of CDKs by CAK (Aprelikova et al, 1995), may contribute to the complex network of cell-cycle control (Clarke, 1995;Hunter and Pines, 1994;Morgan, 1995;Sherr and Roberts, 1995).…”

“…To ensure proper timing and mutual coordination of the major cell-cycle events, the CDKs are carefully regulated by multiple mechanisms (Clarke, 1995;Doree and Galas, 1994;Morgan, 1995;Sherr and Roberts, 1995). One such mechanism is an activatory phosphorylation of a conserved threonine residue (Thr 160 in CDK2, Thr 161 in CDC2 and their equivalents in other CDKs) within the T loop of various CDKs (Clarke, 1995;Doree and Galas, 1994;Morgan, 1995). This phosphorylation appears to alter conformation of the T loop, to stabilize the CDK-cyclin interaction and probably to render the catalytic domain of CDKs accessible to their cellular protein substrates Pines, 1995).…”

“…A kinase responsible for phosphorylating the threonine residue has been identified in various organisms and designated CAK, for CDK-activating kinase (Darbon et al, 1994; Fesquet etal., 1993 (Shuttleworth et al, 1990). The catalytic subunit of CAK itself, therefore, appears to be a member of the CDK family, and it requires binding to a regulatory cyclin subunit to become active (Clarke, 1995;Fisher and Morgan, 1994;Makela et al, 1994;Morgan, 1995). The CAK has therefore been recently renamed CDK7 and its novel cyclin partner termed cyclin H Makela et al, 1994).…”

“…CAK purified from frog, starfish, mouse and human cells was shown to contain a 40-kDa catalytic subunit (Darbon et al, 1994; Fesquet et a/., 1993; Matsuoka et a/,, 1994; Poon et a/., 1993; Solomon etal., 1993;Tassan eta/., 1994), previously identified in a genetic screen for new homologues of CDC2 and termed M015 (Shuttleworth et al, 1990). The catalytic subunit of CAK itself, therefore, appears to be a member of the CDK family, and it requires binding to a regulatory cyclin subunit to become active (Clarke, 1995;Fisher and Morgan, 1994;Makela et al, 1994;Morgan, 1995). The CAK has therefore been recently renamed CDK7 and its novel cyclin partner termed cyclin H Makela et al, 1994).…”