Regulation of the localization and stability of Cdc6 in living yeast cells
Regulation of the localization and stability of Cdc6 in living yeast cells
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DOI：
10.1016/s0006-291x(03)01082-9
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发表时间：
2003-07-11
影响因子：
3.1
通讯作者：
Campbell, JL
中科院分区：
文献类型：
--
作者：
Luo, KQ;Elsasser, S;Campbell, JL
Cdc 6蛋白是启动DNA复制的重要调节因子。在G1/S转换之后，Cdc 6通过泛素介导的蛋白水解途径降解。在这项研究中，我们标记的Cdc 6与绿色荧光蛋白（GFP）和使用位点特异性突变，研究Cdc 6的本地化和降解的调节活酵母细胞。我们的主要发现是：（1）Cdc 6-GFP在细胞周期各阶段均主要分布在细胞核内，在G2/M期细胞胞质定位略有增加。(2)这种核定位对于Cdc 6降解至关重要。当N端核定位信号（NLS）突变时。Cdc 6-GFP不再在细胞核中积累，并且突变体cdc 6与野生型相比稳定。(3)推定的CDK磷酸化位点不需要Cdc 6核定位，但对蛋白质稳定性很重要。这些观察结果表明，Cdc 6蛋白的稳定性是由两个因素：核定位和磷酸化的CDK 1。(C)2003 Elsevier Science（美国）。All rights reserved.
The Cdc6 protein is an essential regulator for initiation of DNA replication. Following the G1/S transition, Cdc6 is degraded through a ubiquitin-mediated proteolysis pathway. In this study, we tagged Cdc6 with green fluorescent protein (GFP) and used site-specific mutations to study the regulation of Cdc6 localization and degradation in living yeast cells. Our major findings are: (1) Cdc6-GFP distributes predominantly in the nucleus in all cell cycle stages, with a small increase in cytoplasmic localization in G2/M cells. (2) This nuclear localization is critical for Cdc6 degradation. When the N-terminal nuclear localization signal (NLS) was mutated. Cdc6-GFP no longer accumulated in the nucleus, and the mutant cdc6 was stabilized compared to wild type. (3) The putative CDK phosphorylation sites are not required for Cdc6 nuclear localization, but are important for protein stability. These observations suggest that the stability of Cdc6 protein is regulated by two factors: nuclear localization and phosphorylation by CDK1. (C) 2003 Elsevier Science (USA). All rights reserved.