Department of Molecular Cellular and Biochemistry, The Ohio State Comprehensive Cancer Center, The Ohio State University, Columbus, OH, USA.
Cell Cycle. 2012 Aug 1;11(15):2828-32. doi: 10.4161/cc.21168.
The origin recognition complex 1 (ORC1) is the largest subunit of the ORC, the heteromeric hexamer. ORC1 is an essential component of the pre-replicative complex (pre-RC) that licenses eukaryote DNA replication origins. The levels of ORC1 fluctuate during the mitotic cell cycle in Drosophila as well as in some human cells. Proteolysis of ORC1 occurs at the end of M phase in Drosophila, which is mediated by the anaphase-promoting complex (APC), and in late S phase in human cells by Skip-Cullin-F box (SCF). Previously we showed that proteolysis of ORC1 by APC is mediated by the ORC1 destruction box (the O-box), an APC motif conserved among species yet distinct from the D-box or KEN-box. Recently we showed that replacing the O-box with the D-box (ORC1O→D) changes the degradation profile of ORC1 during a canonical cell cycle. Here we report further characterization of the ORC1O→D allele that turned out to be a useful tool to examine the function of ORC1 in other modes of DNA replication during oogenesis. In endoreplication stages ORC1O→D does not change any DNA content profiles, consistent with our previous finding that ORC is dispensable for endoreplication. However, in amplification stage replication efficiency of ORC1O→D is drastically reduced, which resulted in amplification defects that led to thin egg shell phenotype. Taken together, our analyses show that orc1 allele newly identified is female sterile and possesses a unique feature of phenotypes that are distinct in different modes of DNA replication.
起始识别复合物 1（ORC1）是 ORC 的最大亚基，异源六聚体。ORC1 是复制前复合物（pre-RC）的一个必需组成部分，该复合物使真核生物 DNA 复制起始。在果蝇的有丝分裂细胞周期以及一些人类细胞中，ORC1 的水平会发生波动。在果蝇中，ORC1 的蛋白水解作用发生在 M 期结束时，由后期促进复合物（APC）介导，而在人类细胞中，在 S 期后期由 Skip-Cullin-F -box（SCF）介导。此前我们表明，APC 介导的 ORC1 蛋白水解作用由 ORC1 破坏盒（O 盒）介导，该盒是物种间保守的 APC 基序，但与 D 盒或 KEN 盒不同。最近我们表明，用 D 盒（ORC1O→D）取代 O 盒会改变 ORC1 在典型细胞周期中的降解谱。在这里，我们进一步描述了 ORC1O→D 等位基因的特征，该等位基因是在卵发生过程中研究 ORC1 在其他 DNA 复制模式中的功能的有用工具。在复制扩增阶段，ORC1O→D 不会改变任何 DNA 含量谱，这与我们之前的发现一致，即 ORC 对于复制扩增是可有可无的。然而，在扩增阶段，ORC1O→D 的复制效率大大降低，导致扩增缺陷，导致薄蛋壳表型。总之，我们的分析表明，新鉴定的 orc1 等位基因是雌性不育的，并具有在不同 DNA 复制模式中独特的表型特征。
