Related Experiment Videos
Systematic analysis of DNA crosslink repair pathways during development and aging in Caenorhabditis elegans
David M Wilson1, Matthias Rieckher2, Ashley B Williams2
1Laboratory of Molecular Gerontology, National Institute on Aging, Intramural Research Program, National Institutes of Health, Baltimore, MD 21224, USA.
Abstract:
DNA interstrand crosslinks (ICLs) are generated by endogenous sources and chemotherapeutics, and pose a threat to genome stability and cell survival. Using Caenorhabditis elegans mutants, we identify DNA repair factors that protect against the genotoxicity of ICLs generated by trioxsalen/ultraviolet A (TMP/UVA) during development and aging. Mutations in nucleotide excision repair (NER) components (e.g. XPA-1 and XPF-1) imparted extreme sensitivity to TMP/UVA relative to wild-type animals, manifested as developmental arrest, defects in adult tissue morphology and functionality, and shortened lifespan. Compensatory roles for global-genome (XPC-1) and transcription-coupled (CSB-1) NER in ICL sensing were exposed. The analysis also revealed contributions of homologous recombination (BRC-1/BRCA1), the MUS-81, EXO-1, SLX-1 and FAN-1 nucleases, and the DOG-1 (FANCJ) helicase in ICL resolution, influenced by the replicative-status of the cell/tissue. No obvious or critical role in ICL repair was seen for non-homologous end-joining (cku-80) or base excision repair (nth-1, exo-3), the Fanconi-related proteins BRC-2 (BRCA2/FANCD1) and FCD-2 (FANCD2), the WRN-1 or HIM-6 (BLM) helicases, or the GEN-1 or MRT-1 (SNM1) nucleases. Our efforts uncover replication-dependent and -independent ICL repair networks, and establish nematodes as a model for investigating the repair and consequences of DNA crosslinks in metazoan development and in adult post-mitotic and proliferative germ cells.
Insights
DNA interstrand crosslinks (ICLs) threaten genome stability. Using C. elegans, we identified key DNA repair factors, including nucleotide excision repair (NER) and homologous recombination, crucial for repairing ICLs and ensuring cell survival during development and aging.
Area of Science:
- Genetics and Molecular Biology
- DNA Repair Mechanisms
- Developmental Biology
Background:
- DNA interstrand crosslinks (ICLs) are DNA lesions arising from endogenous processes and chemotherapy, posing significant risks to genome integrity and cellular viability.
- Understanding the pathways that repair ICLs is crucial for mitigating genotoxicity and its consequences on development and aging.
Purpose of the Study:
- To identify DNA repair factors that protect against ICL genotoxicity in a metazoan model system, Caenorhabditis elegans.
- To elucidate the roles of various DNA repair pathways, including nucleotide excision repair (NER) and homologous recombination, in response to ICLs during development and aging.
Main Methods:
- Utilized Caenorhabditis elegans mutants deficient in specific DNA repair genes.
- Exposed animals to trioxsalen/ultraviolet A (TMP/UVA) to induce ICLs.
- Assessed developmental progression, tissue morphology, functionality, and lifespan as indicators of genotoxicity and repair efficacy.
Main Results:
- Mutations in NER components (XPA-1, XPF-1) caused extreme sensitivity to TMP/UVA, leading to developmental arrest, tissue defects, and reduced lifespan.
- Identified compensatory roles for global-genome NER (XPC-1) and transcription-coupled NER (CSB-1) in ICL sensing.
- Revealed contributions of homologous recombination (BRC-1/BRCA1), specific nucleases (MUS-81, EXO-1, SLX-1, FAN-1), and the DOG-1 (FANCJ) helicase in ICL resolution, dependent on replication status.
- Demonstrated minimal roles for non-homologous end-joining, base excision repair, Fanconi anemia pathway proteins (BRC-2, FCD-2), and certain helicases (WRN-1, HIM-6) and nucleases (GEN-1, MRT-1) in ICL repair.
Conclusions:
- Uncovered distinct replication-dependent and -independent DNA repair networks for ICLs.
- Established Caenorhabditis elegans as a valuable model for studying DNA crosslink repair and its impact on metazoan development and adult tissues.