Moorthamer M, Panchal M, Greenhalf W, Chaudhuri B
Oncology Research, Novartis Pharma AG, Basel, Switzerland.
Biochem Biophys Res Commun. 1998 Sep 29;250(3):791-7. doi: 10.1006/bbrc.1998.9348.
Cyclin-dependent kinase 4 (Cdk4) activity is misregulated in most cancers. Loss of Cdk4 regulation can occur through overexpression of Cdk4 catalytic subunit or its regulatory partner cyclin D1, or if the Cdk4-specific inhibitory protein p16(INK4A) is inactive. We have attempted to express the two human subunits, Cdk4 and cyclin D1, in the yeast Saccharomyces cerevisiae. Surprisingly, expression of Cdk4 alone, under control of the strong GAL promoter, inhibits cell growth. Coexpression of both subunits allows formation of an active Cdk4-cyclin D1 complex which accentuates growth arrest. In cells expressing Cdk4 only, growth is restored by overexpressing human Cdc37, a Cdk4-binding molecular chaperone. Interestingly, the effect of Cdk4 on yeast is also overcome by both p16- and p21-families of Cdk-inhibitory proteins. Moreover, flavopiridol, a compound which inhibits Cdk4 enzyme activity, restores cell division. The fact that p16(INK4A) and flavopiridol negate Cdk4-mediated suppression of yeast cell growth implies that this simple system can be used as a screen for identifying Cdk4-specific antagonists which may mimic p16(INK4A) in the cancer cell cycle.
细胞周期蛋白依赖性激酶4（Cdk4）的活性在大多数癌症中存在失调。Cdk4调控的丧失可通过Cdk4催化亚基或其调节伴侣细胞周期蛋白D1的过表达而发生，或者如果Cdk4特异性抑制蛋白p16（INK4A）无活性。我们试图在酿酒酵母中表达两个人类亚基，即Cdk4和细胞周期蛋白D1。令人惊讶的是，在强GAL启动子的控制下，单独表达Cdk4会抑制细胞生长。两个亚基的共表达允许形成活性Cdk4-细胞周期蛋白D1复合物，这加剧了生长停滞。在仅表达Cdk4的细胞中，通过过表达人Cdc37（一种Cdk4结合分子伴侣）可恢复生长。有趣的是，Cdk4对酵母的影响也被Cdk抑制蛋白的p16和p21家族所克服。此外，一种抑制Cdk4酶活性的化合物黄酮哌啶醇可恢复细胞分裂。p16（INK4A）和黄酮哌啶醇消除Cdk4介导的酵母细胞生长抑制这一事实表明，这个简单的系统可用于筛选识别可能在癌细胞周期中模拟p16（INK4A）的Cdk4特异性拮抗剂。
