Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator.
Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator.
复制标题
古细菌整体 DNA 损伤响应调节因子 Orc1-2 功能域的剖析
DOI：
10.3390/ijms232314609
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发表时间：
2022-11-23
影响因子：
5.6
通讯作者：
中科院分区：
文献类型：
--
作者：
Orc 1 -2是古细菌/真核生物Orc 1蛋白的非起始同源物，其在DNA损伤响应（DDR）表达中起全局调节剂的作用。对于Orc 1启动子，DDR调节子含有AAA+ ATP酶结构域、启动子特异性基序（ISM）和翼形螺旋（wH）DNA结合结构域，它们也以类似的方式组织。为了研究Orc 1 -2如何介导DDR调节，用Islandicus构建了使这些功能结构域中的每一个失活的Orc 1 -2突变体，并对其进行了遗传表征。我们发现，每个功能域的中断完全取消了DDR的调节，在这些orc 1 -2突变体。引人注目的是，Orc 1 -2的ATP水解的失活使得突变体不能存活。然而，细胞致死性可以通过相同蛋白中DNA结合的缺陷来抑制，并且其发生不依赖于任何DNA损伤信号。然后获得突变体Orc 1 -2蛋白，并在体外研究DNA结合。这表明AAA+ ATP酶和wH结构域都参与DNA结合，其中wH中的ISM和R381 R383负责特异性DNA结合。我们进一步表明，Orc 1 -2的调节发生在两个不同的步骤：（a）引发细胞分裂抑制在低Orc 1 -2含量，这种调节是由ATP结合和关闭ATP水解打开，在关闭调节的任何故障导致生长抑制和细胞死亡;（B）激活DDR基因编码的DNA修复蛋白的表达在Orc 1 -2的水平升高。
Orc1-2 is a non-initiator ortholog of archaeal/eukaryotic Orc1 proteins, which functions as a global regulator in DNA damage-responsive (DDR) expression. As for Orc1 initiators, the DDR regulator harbors an AAA+ ATPase domain, an Initiator-Specific Motif (ISM) and a winged-helix (wH) DNA-binding domain, which are also organized in a similar fashion. To investigate how Orc1-2 mediates the DDR regulation, the orc1-2 mutants inactivating each of these functional domains were constructed with Saccharolobus islandicus and genetically characterized. We found that disruption of each functional domain completely abolished the DDR regulation in these orc1-2 mutants. Strikingly, inactivation of ATP hydrolysis of Orc1-2 rendered an inviable mutant. However, the cell lethality can be suppressed by the deficiency of the DNA binding in the same protein, and it occurs independent of any DNA damage signal. Mutant Orc1-2 proteins were then obtained and investigated for DNA-binding in vitro. This revealed that both the AAA+ ATPase and the wH domains are involved in DNA-binding, where ISM and R381R383 in wH are responsible for specific DNA binding. We further show that Orc1-2 regulation occurs in two distinct steps: (a) eliciting cell division inhibition at a low Orc1-2 content, and this regulation is switched on by ATP binding and turned off by ATP hydrolysis; any failure in turning off the regulation leads to growth inhibition and cell death; (b) activation of the expression of DDR gene encoding DNA repair proteins at an elevated level of Orc1-2.
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影响因子：
2.9
作者：
De Felice, M;Esposito, L;Pisani, FM
通讯作者：
Pisani, FM
DOI：
10.2174/1874091x00802010129
发表时间：
2008
期刊：
The open biochemistry journal
影响因子：
--
作者：
Haugland, Gyri Teien;Innselset, Marte;Madern, Dominique;Birkeland, Nils-Kare
通讯作者：
Birkeland, Nils-Kare
影响因子：
10.5
作者：
Greci MD;Bell SD
通讯作者：
Bell SD
影响因子：
5.6
作者：
Han W;Feng X;She Q
通讯作者：
She Q
影响因子：
3.2
作者：
Grabowski, B;Kelman, Z
通讯作者：
Kelman, Z