Yahyaoui Wafaa, Callejo Mario, Price Gerald B, Zannis-Hadjopoulos Maria
McGill Cancer Centre, Montreal, Quebec, Canada.
BMC Mol Biol. 2007 Apr 12;8:27. doi: 10.1186/1471-2199-8-27.
Initiation of eukaryotic DNA replication involves many protein-protein and protein-DNA interactions. We have previously shown that 14-3-3 proteins bind cruciform DNA and associate with mammalian and yeast replication origins in a cell cycle dependent manner.
By expressing the human 14-3-3epsilon, as the sole member of 14-3-3 proteins family in Saccharomyces cerevisiae, we show that 14-3-3epsilon complements the S. cerevisiae Bmh1/Bmh2 double knockout, conserves its cruciform binding activity, and associates in vivo with the yeast replication origins ARS307. Deletion of the alpha5-helix, the potential cruciform binding domain of 14-3-3, decreased the cruciform binding activity of the protein as well as its association with the yeast replication origins ARS307 and ARS1. Furthermore, the mutant cells had a reduced ability to stably maintain plasmids bearing one or multiple origins.
14-3-3, a cruciform DNA binding protein, associates with yeast origins of replication and functions as an initiator of DNA replication, presumably through binding to cruciform DNA forming at yeast replicators.
真核生物DNA复制的起始涉及许多蛋白质-蛋白质和蛋白质-DNA相互作用。我们之前已经表明，14-3-3蛋白结合十字形DNA，并以细胞周期依赖性方式与哺乳动物和酵母的复制起点相关联。
通过在酿酒酵母中表达人14-3-3ε作为14-3-3蛋白家族的唯一成员，我们表明14-3-3ε补充了酿酒酵母Bmh1/Bmh2双敲除，保留了其十字形结合活性，并在体内与酵母复制起点ARS307相关联。删除14-3-3的潜在十字形结合结构域α5螺旋，降低了该蛋白的十字形结合活性以及其与酵母复制起点ARS307和ARS1的关联。此外，突变细胞稳定维持携带一个或多个起点的质粒的能力降低。
14-3-3，一种十字形DNA结合蛋白，与酵母复制起点相关联，并作为DNA复制的起始因子发挥作用，可能是通过结合在酵母复制子处形成的十字形DNA。
