Smeets Monique F M A, Francesconi Stefania, Baldacci Giuseppe
UMR2027, Génotoxicologie et Cycle Cellulaire, Institut Curie, 91405 Orsay, France.
Genes Cells. 2003 Jul;8(7):573-86. doi: 10.1046/j.1365-2443.2003.00657.x.
In eukaryotic cells DNA structure checkpoints organize the cellular responses of DNA repair and transient cell cycle arrest and thereby ensure genomic stability. To investigate the exact role of crb2+ in the DNA damage checkpoint response, a genetic screen was carried out in order to identify suppressors of the conditional MMS sensitivity of a crb2-1 mutant. Here we report the isolation of rhp51+ as a multicopy suppressor.
We show that suppression is not specific for the checkpoint mutant while it is specific for the MMS treatment. Rescue by rhp51+ over-expression is not a consequence of increased recombination repair or checkpoint compensation and epistasis analysis confirms that crb2+ and rhp51+ function in different pathways. A tight linkage between the two pathways is nevertheless suggested by the complementary expression or modification of Crb2 and Rhp51 proteins. Crb2 protein stability is down-regulated when Rhp51 is over-expressed and up-regulated in the absence of Rhp51. The up-regulation of Crb2 is independent of the activation of DNA structure checkpoints. Conversely Rhp51 is more readily activated and differentially modified in the absence of Crb2 or other checkpoint proteins.
We conclude that fission yeast Crb2 and Rhp51 function in two parallel, tightly connected and coordinately regulated pathways.
在真核细胞中，DNA结构检查点可组织细胞对DNA修复和短暂细胞周期停滞的反应，从而确保基因组稳定性。为了研究crb2+在DNA损伤检查点反应中的具体作用，进行了一项遗传筛选，以鉴定crb2-1突变体条件性甲磺酸甲酯（MMS）敏感性的抑制因子。在此，我们报告分离出rhp51+作为多拷贝抑制因子。
我们发现这种抑制并非针对检查点突变体具有特异性，而是对MMS处理具有特异性。rhp51+过表达的拯救作用并非增加重组修复或检查点补偿的结果，上位性分析证实crb2+和rhp51+在不同途径中发挥作用。然而，Crb2和Rhp51蛋白的互补表达或修饰表明这两条途径之间存在紧密联系。当Rhp51过表达时，Crb2蛋白稳定性下调，而在没有Rhp51时则上调。Crb2的上调与DNA结构检查点的激活无关。相反，在没有Crb2或其他检查点蛋白的情况下，Rhp51更容易被激活并发生差异修饰。
我们得出结论，裂殖酵母Crb2和Rhp51在两条平行、紧密连接且协调调节的途径中发挥作用。
