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The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination
DNA Ligase IV is responsible for the repair of DNA double-strand breaks (DSB), including DSBs that are generated during V(D)J recombination. Like other DNA ligases, Ligase IV contains a catalytic core with three subdomains—the DNA binding (DBD), the nucleotidyltransferase (NTD), and the oligonucleot...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Public Library of Science
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9956705/ https://www.ncbi.nlm.nih.gov/pubmed/36827388 http://dx.doi.org/10.1371/journal.pone.0282236 |
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author | Malashetty, Vidyasagar Au, Audrey Chavez, Jose Hanna, Mary Chu, Jennifer Penna, Jesse Cortes, Patricia |
author_facet | Malashetty, Vidyasagar Au, Audrey Chavez, Jose Hanna, Mary Chu, Jennifer Penna, Jesse Cortes, Patricia |
author_sort | Malashetty, Vidyasagar |
collection | PubMed |
description | DNA Ligase IV is responsible for the repair of DNA double-strand breaks (DSB), including DSBs that are generated during V(D)J recombination. Like other DNA ligases, Ligase IV contains a catalytic core with three subdomains—the DNA binding (DBD), the nucleotidyltransferase (NTD), and the oligonucleotide/oligosaccharide-fold subdomain (OBD). Ligase IV also has a unique C-terminal region that includes two BRCT domains, a nuclear localization signal sequence and a stretch of amino acid that participate in its interaction with XRCC4. Out of the three mammalian ligases, Ligase IV is the only ligase that participates in and is required for V(D)J recombination. Identification of the minimal domains within DNA Ligase IV that contribute to V(D)J recombination has remained unresolved. The interaction of the Ligase IV DNA binding domain with Artemis, and the interaction of its C-terminal region with XRCC4, suggest that both of these regions that also interact with the Ku70/80 heterodimer are important and might be sufficient for mediating participation of DNA Ligase IV in V(D)J recombination. This hypothesis was investigated by generating chimeric ligase proteins by swapping domains, and testing their ability to rescue V(D)J recombination in Ligase IV-deficient cells. We demonstrate that a fusion protein containing Ligase I NTD and OBDs flanked by DNA Ligase IV DBD and C-terminal region is sufficient to support V(D)J recombination. This chimeric protein, which we named Ligase 37, complemented formation of coding and signal joints. Coding joints generated with Ligase 37 were shorter than those observed with wild type DNA Ligase IV. The shorter length was due to increased nucleotide deletions and decreased nucleotide insertions. Additionally, overexpression of Ligase 37 in a mouse pro-B cell line supported a shift towards shorter coding joints. Our findings demonstrate that the ability of DNA Ligase IV to participate in V(D)J recombination is in large part mediated by its DBD and C-terminal region. |
format | Online Article Text |
id | pubmed-9956705 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-99567052023-02-25 The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination Malashetty, Vidyasagar Au, Audrey Chavez, Jose Hanna, Mary Chu, Jennifer Penna, Jesse Cortes, Patricia PLoS One Research Article DNA Ligase IV is responsible for the repair of DNA double-strand breaks (DSB), including DSBs that are generated during V(D)J recombination. Like other DNA ligases, Ligase IV contains a catalytic core with three subdomains—the DNA binding (DBD), the nucleotidyltransferase (NTD), and the oligonucleotide/oligosaccharide-fold subdomain (OBD). Ligase IV also has a unique C-terminal region that includes two BRCT domains, a nuclear localization signal sequence and a stretch of amino acid that participate in its interaction with XRCC4. Out of the three mammalian ligases, Ligase IV is the only ligase that participates in and is required for V(D)J recombination. Identification of the minimal domains within DNA Ligase IV that contribute to V(D)J recombination has remained unresolved. The interaction of the Ligase IV DNA binding domain with Artemis, and the interaction of its C-terminal region with XRCC4, suggest that both of these regions that also interact with the Ku70/80 heterodimer are important and might be sufficient for mediating participation of DNA Ligase IV in V(D)J recombination. This hypothesis was investigated by generating chimeric ligase proteins by swapping domains, and testing their ability to rescue V(D)J recombination in Ligase IV-deficient cells. We demonstrate that a fusion protein containing Ligase I NTD and OBDs flanked by DNA Ligase IV DBD and C-terminal region is sufficient to support V(D)J recombination. This chimeric protein, which we named Ligase 37, complemented formation of coding and signal joints. Coding joints generated with Ligase 37 were shorter than those observed with wild type DNA Ligase IV. The shorter length was due to increased nucleotide deletions and decreased nucleotide insertions. Additionally, overexpression of Ligase 37 in a mouse pro-B cell line supported a shift towards shorter coding joints. Our findings demonstrate that the ability of DNA Ligase IV to participate in V(D)J recombination is in large part mediated by its DBD and C-terminal region. Public Library of Science 2023-02-24 /pmc/articles/PMC9956705/ /pubmed/36827388 http://dx.doi.org/10.1371/journal.pone.0282236 Text en © 2023 Malashetty et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Malashetty, Vidyasagar Au, Audrey Chavez, Jose Hanna, Mary Chu, Jennifer Penna, Jesse Cortes, Patricia The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination |
title | The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination |
title_full | The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination |
title_fullStr | The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination |
title_full_unstemmed | The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination |
title_short | The DNA binding domain and the C-terminal region of DNA Ligase IV specify its role in V(D)J recombination |
title_sort | dna binding domain and the c-terminal region of dna ligase iv specify its role in v(d)j recombination |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9956705/ https://www.ncbi.nlm.nih.gov/pubmed/36827388 http://dx.doi.org/10.1371/journal.pone.0282236 |
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