Martinez Juan S, Jeong Dah-Eun, Choi Eunyoung, Billings Brian M, Hall Mark C
Department of Biochemistry, Purdue University, West Lafayette, IN, USA.
Mol Cell Biol. 2006 Dec;26(24):9162-76. doi: 10.1128/MCB.00603-06. Epub 2006 Oct 9.
Cdh1 is a coactivator of the anaphase-promoting complex/cyclosome (APC/C) and contributes to mitotic exit and G1 maintenance by facilitating the polyubiquitination and subsequent proteolysis of specific substrates. Here, we report that budding yeast Cdh1 is a component of a cell cycle-regulated complex that includes the 14-3-3 homologs Bmh1 and Bmh2 and a previously uncharacterized protein, which we name Acm1 (APC/CCdh1 modulator 1). Association of Cdh1 with Bmh1 and Bmh2 requires Acm1, and the Acm1 protein is cell cycle regulated, appearing late in G1 and disappearing in late M. In acm1Delta strains, Cdh1 localization to the bud neck and association with two substrates, Clb2 and Hsl1, were strongly enhanced. Several lines of evidence suggest that Acm1 can suppress APC/CCdh1-mediated proteolysis of mitotic cyclins. First, overexpression of Acm1 fully restored viability to cells expressing toxic levels of Cdh1 or a constitutively active Cdh1 mutant lacking inhibitory phosphorylation sites. Second, overexpression of Acm1 was toxic in sic1Delta cells. Third, ACM1 deletion exacerbated a low-penetrance elongated-bud phenotype caused by modest overexpression of Cdh1. This bud elongation was independent of the morphogenesis checkpoint, and the combination of acm1Delta and hsl1Delta resulted in a dramatic enhancement of bud elongation and G2/M delay. Effects on bud elongation were attenuated when Cdh1 was replaced with a mutant lacking the C-terminal IR dipeptide, suggesting that APC/C-dependent proteolysis is required for this phenotype. We propose that Acm1 and Bmh1/Bmh2 constitute a specialized inhibitor of APC/CCdh1.
Cdh1是后期促进复合物/细胞周期体（APC/C）的一种共激活因子，通过促进特定底物的多聚泛素化及随后的蛋白水解作用，对有丝分裂退出和G1期维持发挥作用。在此，我们报告出芽酵母Cdh1是一个细胞周期调控复合物的组成成分，该复合物包括14-3-3同源物Bmh1和Bmh2以及一种先前未被鉴定的蛋白质，我们将其命名为Acm1（APC/CCdh1调节因子1）。Cdh1与Bmh1和Bmh2的结合需要Acm1，并且Acm1蛋白受细胞周期调控，在G1期晚期出现，在M期晚期消失。在acm1Δ菌株中，Cdh1定位于芽颈以及与两种底物Clb2和Hsl1的结合显著增强。多项证据表明，Acm1可以抑制APC/CCdh1介导的有丝分裂周期蛋白的蛋白水解作用。首先，Acm1的过表达完全恢复了表达毒性水平Cdh1或缺乏抑制性磷酸化位点的组成型活性Cdh1突变体的细胞的活力。其次，Acm1的过表达在sic1Δ细胞中具有毒性。第三，ACM1缺失加剧了由Cdh1适度过表达引起的低频率延长芽表型。这种芽伸长与形态发生检查点无关，并且acm1Δ和hsl1Δ的组合导致芽伸长显著增强和G2/M期延迟。当用缺乏C末端IR二肽的突变体替换Cdh1时，对芽伸长的影响减弱，这表明该表型需要APC/C依赖性蛋白水解作用。我们提出Acm1和Bmh1/Bmh2构成了APC/CCdh1的一种特殊抑制剂。
