Inhibition of cdk2 Activating Phosphorylation by Mevastatin

Chinweike Ukomadu,Anindya Dutta

doi:10.1074/jbc.m208658200

Chinweike Ukomadu, Anindya Dutta

Open Access

https://doi.org/10.1074/jbc.m208658200

Copy DOI

Abstract

Phosphorylation of cdk2 on threonine 160 is essential for kinase activity. Mevastatin, an inhibitor of cholesterol synthesis, inhibits cell growth through inhibition of cdk2 and this has been suggested to be due to enhancement of p21 levels. In a prostate cancer cell line, PC3, mevastatin treatment led to elevated levels of p21 and caused a small increase in the p21 associated with cdk2. However, this increase in the associated p21 appeared out of proportion with the resulting dramatic inhibition of kinase activity. Using RNA interference we show that mevastatin inhibits cdk2 activity despite lack of induction of p21, p27, and p57. Instead the kinase was inhibited due to a decrease in activating phosphorylation. Phosphorylation of cdk2 from mevastatin-treated cells with exogenous cyclin-dependent kinase (cdk)-activating enzymes restored its functional activity. The only known mammalian cyclin H.cdk7.mat1 complex (cdk2-activating kinase, Cak), was not inhibited by mevastatin, suggesting either that a different CAK is responsible for cdk2 phosphorylation in vivo or that the regulation is at the level of substrate accessibility or of cdk2 dephosphorylation. These results suggest that mevastatin inhibits cdk2 activity in PC3 cells through the inhibition of Thr-160 phosphorylation of cdk2, providing a novel example of regulation of cdk2 at this level.

Highlights

The basic cell cycle machinery is composed of regulatory cyclin subunits complexed to cyclin-dependent kinase1 subunits [1]
Mevastatin Treatment Leads to a G1 Block and Decrease in Kinase Activity of cdk2—As has been reported with lovastatin treatment of PC3-M cells [17], mevastatin treatment leads to a significant decrease in the S phase population with a concurrent increase in G1 noted by 24 h, and a more pronounced G1 block by 36 h (Fig. 1A)
There is no doubt that p21 levels are elevated in mevastatin-treated cells, our studies suggest that p21 may not always be the active inhibitor of the cdk2 kinase

Summary

Introduction

The basic cell cycle machinery is composed of regulatory cyclin subunits complexed to cyclin-dependent kinase (cdk) subunits [1]. The cyclin1⁄7cdk complex phosphorylates specific substrates in a cell cycle-dependent manner. The activation of cyclin E or A1⁄7cdk requires the dephosphorylation of Thr-14 and Tyr-15 and the phosphorylation of Thr-160. Regulation of cdk in response to different stimuli has until now been described at the level of CKI induction or changes in the inhibitory phosphorylation on Thr-14 and Tyr-15. Known as “statins,” these drugs are used for treatment of hypercholesterolemia [10] One explanation for their growth-altering effects is that the lack of isoprenylation of key regulatory proteins like Ras, Rap, and others leads to defective subcellular localization of these key proteins. The increased association of p21 with cyclin1⁄7cdk that accompanies the cdk kinase inhibition is believed to account for the cell cycle block. Instead a decrease in the activating phosphorylation of cdk likely accounts for the antiproliferative effects of this drug

Methods

Results

Conclusion