Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
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Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator. / Liu, Xiaotong; Sun, Mengmeng; Xu, Ruyi; Shen, Yulong; Huang, Qihong; Feng, Xu; She, Qunxin.
In: International Journal of Molecular Sciences, Vol. 23, No. 23, 14609, 2022.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
AU - Liu, Xiaotong
AU - Sun, Mengmeng
AU - Xu, Ruyi
AU - Shen, Yulong
AU - Huang, Qihong
AU - Feng, Xu
AU - She, Qunxin
N1 - Publisher Copyright: © 2022 by the authors.
PY - 2022
Y1 - 2022
N2 - 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.
AB - 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.
KW - AAA+ ATPase
KW - archaea
KW - DNA damage response
KW - global regulator
KW - origin recognition complex
KW - winged-helix domain
U2 - 10.3390/ijms232314609
DO - 10.3390/ijms232314609
M3 - Journal article
C2 - 36498936
AN - SCOPUS:85143776340
VL - 23
JO - International Journal of Molecular Sciences (Online)
JF - International Journal of Molecular Sciences (Online)
SN - 1661-6596
IS - 23
M1 - 14609
ER -
ID: 330383527