Cargando…

Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator

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 dom...

Descripción completa

Detalles Bibliográficos
Autores principales: Liu, Xiaotong, Sun, Mengmeng, Xu, Ruyi, Shen, Yulong, Huang, Qihong, Feng, Xu, She, Qunxin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9738581/
https://www.ncbi.nlm.nih.gov/pubmed/36498936
http://dx.doi.org/10.3390/ijms232314609
_version_ 1784847581293051904
author Liu, Xiaotong
Sun, Mengmeng
Xu, Ruyi
Shen, Yulong
Huang, Qihong
Feng, Xu
She, Qunxin
author_facet Liu, Xiaotong
Sun, Mengmeng
Xu, Ruyi
Shen, Yulong
Huang, Qihong
Feng, Xu
She, Qunxin
author_sort Liu, Xiaotong
collection PubMed
description 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.
format Online
Article
Text
id pubmed-9738581
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-97385812022-12-11 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 Int J Mol Sci Article 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. MDPI 2022-11-23 /pmc/articles/PMC9738581/ /pubmed/36498936 http://dx.doi.org/10.3390/ijms232314609 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Liu, Xiaotong
Sun, Mengmeng
Xu, Ruyi
Shen, Yulong
Huang, Qihong
Feng, Xu
She, Qunxin
Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
title Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
title_full Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
title_fullStr Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
title_full_unstemmed Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
title_short Dissection of Functional Domains of Orc1-2, the Archaeal Global DNA Damage-Responsive Regulator
title_sort dissection of functional domains of orc1-2, the archaeal global dna damage-responsive regulator
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9738581/
https://www.ncbi.nlm.nih.gov/pubmed/36498936
http://dx.doi.org/10.3390/ijms232314609
work_keys_str_mv AT liuxiaotong dissectionoffunctionaldomainsoforc12thearchaealglobaldnadamageresponsiveregulator
AT sunmengmeng dissectionoffunctionaldomainsoforc12thearchaealglobaldnadamageresponsiveregulator
AT xuruyi dissectionoffunctionaldomainsoforc12thearchaealglobaldnadamageresponsiveregulator
AT shenyulong dissectionoffunctionaldomainsoforc12thearchaealglobaldnadamageresponsiveregulator
AT huangqihong dissectionoffunctionaldomainsoforc12thearchaealglobaldnadamageresponsiveregulator
AT fengxu dissectionoffunctionaldomainsoforc12thearchaealglobaldnadamageresponsiveregulator
AT shequnxin dissectionoffunctionaldomainsoforc12thearchaealglobaldnadamageresponsiveregulator