Recruitment of Rad17-RFC complex to DNA

The Rad17-RFC complex is involved in an early stage of the genotoxic stress response. The major function of the protein complex is to load the Rad9-Hus1-Rad1 (9-1-1) complex onto DNA at sites of damage and/or stalled replication forks. This reaction is conceptually similar to the loading of the PCNA sliding clamp onto DNA by RFC. The association of the Rad17-RFC complex with ssDNA or gapped or primed DNA is significantly stimulated by RPA, but not by the heterologous E. coli SSB. Loading of the human 9-1-1 complex onto such DNA templates is also strongly stimulated by cognate RPA, but not yeast RPA. Although Rad17 and Rad9 are substrates of the ATR kinase activity, loading of the Rad17 and 9-1-1 complexes onto DNA occurs independent of ATR.<p>The Rad17-RFC complex is a heteropentamer structurally similar to RFC. The complex contains the four smaller RFC subunits (Rfc2 [p37], Rfc3 [p36], Rfc4 [p40], and Rfc5 [p38]) and the 75 kDa Rad17 subunit in place of the Rfc1 [p140] subunit. The Rad17 complex contains a weak ATPase that is slightly stimulated by primed DNA. Along with binding the 9-1-1 complex and RPA, the Rad17-RFC complex interacts with human MCM7 protein. Each of these interactions is critical for Chk1 activation.<p>The Rad17 subunit is conserved evolutionarily with the protein showing 49% identity at the amino acid level with the S. pombe rad17 protein. Targeted deletion of the N-terminal region of mouse Rad17 leads to embryonic lethality, strongly suggesting that human Rad17 is also essential for long-term viability.<p>Rad17-RFC complex associates with DNA substrates containing ssDNA regions including gapped or primed DNA in an ATP-independent reaction. Loading of the Rad9-Hus1-Rad1 (9-1-1) complex occurs preferentially on DNA substrates containing a 5' recessed end. This contrasts with the loading of PCNA by RFC which preferentially occurs on DNA with 3' recessed ends.

Rad17-RFC复合体在基因毒性应激反应的早期阶段发挥作用。该蛋白质复合体的主要功能是在损伤位点或停滞的复制叉处将Rad9-Hus1-Rad1（9-1-1）复合体加载到DNA上。这一反应在概念上类似于RFC通过PCNA滑动钳将DNA加载的过程。Rad17-RFC复合体与单链DNA、缺口或引物DNA的结合显著受到RPA的刺激，而不受异源的大肠杆菌SSB的影响。将人类9-1-1复合体加载到此类DNA模板上同样受到同源RPA的强烈刺激，而不受酵母RPA的刺激。尽管Rad17和Rad9是ATR激酶活性的底物，但Rad17和9-1-1复合体在DNA上的加载与ATR无关。<p>Rad17-RFC复合体是一个与RFC结构相似的杂五聚体。该复合体包含四个较小的RFC亚基（Rfc2 [p37]、Rfc3 [p36]、Rfc4 [p40]和Rfc5 [p38]）以及取代Rfc1 [p140]亚基的75 kDa的Rad17亚基。Rad17复合体含有一个微弱的ATP酶，该酶在引物DNA的刺激下略有活性。与9-1-1复合体和RPA的结合之外，Rad17-RFC复合体还与人类MCM7蛋白相互作用。这些相互作用对于Chk1的激活至关重要。<p>Rad17亚基在进化上与蛋白质保持一致性，在氨基酸水平上与S. pombe的rad17蛋白具有49%的同源性。小鼠Rad17的N端区域的靶向删除导致胚胎致死，强烈表明人类Rad17对于长期存活也是必不可少的。<p>Rad17-RFC复合体通过与含有单链DNA区域，包括缺口或引物DNA的DNA底物以ATP非依赖性反应相结合。Rad9-Hus1-Rad1（9-1-1）复合体的加载优先发生在含有5' recessed末端的DNA底物上。这与RFC优先加载含有3' recessed末端的DNA的过程形成对比。