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

Abstract

The cyclin-dependent kinase 7 (CDK7) represents the 40-kDa catalytic subunit of the CDK-activating kinase, the enzyme responsible for activatory phosphorylation of multiple CDKs controlling GI, S and G2/M phases of the cell cycle. Here, we surveyed a wide range of normal and tumour cell types, in both cell culture and biopsy specimens, for abundance and subcellular localisation of the CDK7 protein. lmmunoblotting and immunocytochemical analyses showed that CDK7 was (i) ubiquitously expressed in all cell types examined; (ii) exclusively nuclear: (iii) moderately elevated in turnour cells when compared with their normal cell counterparts; (iv) invariant throughout the cell cycle of normal lymphocytes, fibroblasts, breast epithelium and several cancer cell lines; and (v) clearly detectable even in quiescent cells, including highly differentiated cell types in situ. Our data are consistent with the emerging role for CDK7/CAK in multiple biological processes, possibly providing a link between cell-cycle control, transcriptional regulation and genomic integrity control.o 1996 Wiley-Liss, Inc.Transitions through the ordered checkpoints that control the mammalian cell cycle are triggered by activation of a distinct class of serine/threonine kinases, the cyclin-dependent kinases (CDKs). 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).Accumulating evidence implicates deregulation of some of the components of the mammalian cell-cycle machinery in oncogenesis. In particular, over-expression of D-type cyclins or their CDK4 partner kinase appears to be involved in pathogenesis of several human cancer types, and these positive cell cycle regulators qualify as new proto-oncogenes (Bartkova et al., 19956;Hunter and Pines, 1994;Sherr and Roberts, 1995;Strauss et al., 1995). Since CDK7/CAK activates the key CDKs at various cell-cycle transitions, including...

Aberrant p27Kip1 promoter methylation in malignant melanoma

“…The immunoblotting and immunostaining results were con®rmed with two independent antibodies against p27 Kip1 , and parallel examination of a control stable protein, CDK7 (Bartkova et al, 1996), showed comparable abundance and localization in both the melanoma cell lines and normal melanocytes (see Figure 4b for examples). The Kip1 protein in the three melanoma cell lines supports the notion that one allele of the p27…”

Tumor Extension and Cell Proliferation in Adenocarcinomas of the Lung

“…This is in contrast to a previous report in which the proliferative activity in human breast and urinary bladder carcinomas was attributed to up-regulated expression of CAK protein. 38 Assuming that the cyclin A-cdk2 complex is not activated by the positive regulator CAK, one alternative interpretation is that its activity is up-regulated by the suppression of kinase inhibitors. However, our results show that p21 and p27 exhibit higher LIs in the periphery.…”

Cyclins and cyclin-dependent kinases: Comparative study of hepatocellular carcinoma versus cirrhosis

“…In fact, overexpression of cyclin B1, Cdk1, Cdk2, and Cdk4 also has been shown in human cancers other than HCC. [21][22][23][24] Based on these facts, it is of interest to examine other cell cyclerelated proteins, including various cyclins and Cdks, as related to the carcinogenesis or progression in human HCC.…”