Deshaies R J, Chau V, Kirschner M
Department of Biochemistry and Biophysics, University of California Medical Center, San Francisco 94143.
EMBO J. 1995 Jan 16;14(2):303-12. doi: 10.1002/j.1460-2075.1995.tb07004.x.
Recombinant G1 cyclin Cln2p can bind to and stimulate the protein kinase activity of p34CDC28 (Cdc28p) in an extract derived from cyclin-depleted and G1-arrested Saccharomyces cerevisiae cells. Upon activating Cdc28p, Cln2p is extensively phosphorylated and conjugated with multiubiquitin chains. Ubiquitination of Cln2p in vitro requires the Cdc34p ubiquitin-conjugating enzyme, Cdc28p, protein phosphorylation and unidentified factors in yeast extract. Ubiquitination of Cln2p by Cdc34p contributes to the instability of Cln2p in vivo, as the rate of Cln2p degradation is reduced in cdc34ts cells. These results provide a molecular framework for G1 cyclin instability and suggest that a multicomponent, regulated pathway specifies the selective ubiquitination of G1 cyclins.
重组G1细胞周期蛋白Cln2p能够在来自细胞周期蛋白缺失且处于G1期阻滞的酿酒酵母细胞的提取物中，与p34CDC28（Cdc28p）结合并刺激其蛋白激酶活性。激活Cdc28p后，Cln2p会被广泛磷酸化并与多聚泛素链结合。体外Cln2p的泛素化需要Cdc34p泛素结合酶、Cdc28p、蛋白磷酸化以及酵母提取物中的未知因子。Cdc34p介导的Cln2p泛素化在体内导致Cln2p不稳定，因为在cdc34ts细胞中Cln2p的降解速率降低。这些结果为G1细胞周期蛋白的不稳定性提供了分子框架，并表明一条多组分、受调控的途径决定了G1细胞周期蛋白的选择性泛素化。
