Cargando…
The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks
DNA double-strand breaks (DSBs) are a particularly genotoxic type of DNA damage that can result in chromosomal aberrations. Thus, proper repair of DSBs is essential to maintaining genome integrity. DSBs can be repaired by non-homologous end joining (NHEJ), where ends are processed before joining thr...
Autores principales: | , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8468788/ https://www.ncbi.nlm.nih.gov/pubmed/34573412 http://dx.doi.org/10.3390/genes12091430 |
_version_ | 1784573761215791104 |
---|---|
author | Yannuzzi, Ian Butler, Margaret A. Fernandez, Joel LaRocque, Jeannine R. |
author_facet | Yannuzzi, Ian Butler, Margaret A. Fernandez, Joel LaRocque, Jeannine R. |
author_sort | Yannuzzi, Ian |
collection | PubMed |
description | DNA double-strand breaks (DSBs) are a particularly genotoxic type of DNA damage that can result in chromosomal aberrations. Thus, proper repair of DSBs is essential to maintaining genome integrity. DSBs can be repaired by non-homologous end joining (NHEJ), where ends are processed before joining through ligation. Alternatively, DSBs can be repaired through homology-directed repair, either by homologous recombination (HR) or single-strand annealing (SSA). Both types of homology-directed repair are initiated by DNA end resection. In cultured human cells, the protein CtIP has been shown to play a role in DNA end resection through its interactions with CDK, BRCA1, DNA2, and the MRN complex. To elucidate the role of CtIP in a multicellular context, CRISPR/Cas9 genome editing was used to create a DmCtIP(Δ) allele in Drosophila melanogaster. Using the DSB repair reporter assay direct repeat of white (DR-white), a two-fold decrease in HR in DmCtIP(Δ/Δ) mutants was observed when compared to heterozygous controls. However, analysis of HR gene conversion tracts (GCTs) suggests DmCtIP plays a minimal role in determining GCT length. To assess the function of DmCtIP on both short (~550 bp) and long (~3.6 kb) end resection, modified homology-directed SSA repair assays were implemented, resulting in a two-fold decrease in SSA repair in both short and extensive end resection requirements in the DmCtIP(Δ/Δ) mutants compared to heterozygote controls. Through these analyses, we affirmed the importance of end resection on DSB repair pathway choice in multicellular systems, described the function of DmCtIP in short and extensive DNA end resection, and determined the impact of end resection on GCT length during HR. |
format | Online Article Text |
id | pubmed-8468788 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-84687882021-09-27 The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks Yannuzzi, Ian Butler, Margaret A. Fernandez, Joel LaRocque, Jeannine R. Genes (Basel) Article DNA double-strand breaks (DSBs) are a particularly genotoxic type of DNA damage that can result in chromosomal aberrations. Thus, proper repair of DSBs is essential to maintaining genome integrity. DSBs can be repaired by non-homologous end joining (NHEJ), where ends are processed before joining through ligation. Alternatively, DSBs can be repaired through homology-directed repair, either by homologous recombination (HR) or single-strand annealing (SSA). Both types of homology-directed repair are initiated by DNA end resection. In cultured human cells, the protein CtIP has been shown to play a role in DNA end resection through its interactions with CDK, BRCA1, DNA2, and the MRN complex. To elucidate the role of CtIP in a multicellular context, CRISPR/Cas9 genome editing was used to create a DmCtIP(Δ) allele in Drosophila melanogaster. Using the DSB repair reporter assay direct repeat of white (DR-white), a two-fold decrease in HR in DmCtIP(Δ/Δ) mutants was observed when compared to heterozygous controls. However, analysis of HR gene conversion tracts (GCTs) suggests DmCtIP plays a minimal role in determining GCT length. To assess the function of DmCtIP on both short (~550 bp) and long (~3.6 kb) end resection, modified homology-directed SSA repair assays were implemented, resulting in a two-fold decrease in SSA repair in both short and extensive end resection requirements in the DmCtIP(Δ/Δ) mutants compared to heterozygote controls. Through these analyses, we affirmed the importance of end resection on DSB repair pathway choice in multicellular systems, described the function of DmCtIP in short and extensive DNA end resection, and determined the impact of end resection on GCT length during HR. MDPI 2021-09-16 /pmc/articles/PMC8468788/ /pubmed/34573412 http://dx.doi.org/10.3390/genes12091430 Text en © 2021 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 Yannuzzi, Ian Butler, Margaret A. Fernandez, Joel LaRocque, Jeannine R. The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks |
title | The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks |
title_full | The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks |
title_fullStr | The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks |
title_full_unstemmed | The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks |
title_short | The Role of Drosophila CtIP in Homology-Directed Repair of DNA Double-Strand Breaks |
title_sort | role of drosophila ctip in homology-directed repair of dna double-strand breaks |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8468788/ https://www.ncbi.nlm.nih.gov/pubmed/34573412 http://dx.doi.org/10.3390/genes12091430 |
work_keys_str_mv | AT yannuzziian theroleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks AT butlermargareta theroleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks AT fernandezjoel theroleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks AT larocquejeanniner theroleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks AT yannuzziian roleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks AT butlermargareta roleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks AT fernandezjoel roleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks AT larocquejeanniner roleofdrosophilactipinhomologydirectedrepairofdnadoublestrandbreaks |