Mouse Embryonic Fibroblasts Isolated From Nthl1 D227Y Knockin Mice Exhibit Defective DNA Repair and Increased Genome...

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Oxidative DNA damage can lead to genomic instability and diseases like cancer if unrepaired.
• Nth-like DNA glycosylase 1 (NTHL1) is crucial for removing oxidized pyrimidine lesions.
• The NTHL1 D239Y germline variant is found in humans in both heterozygous and homozygous states.

Purpose of the Study:

• To investigate the cellular effects of the NTHL1 D227Y variant (mouse homologue of D239Y) in heterozygous and homozygous states.
• To understand the impact of this variant on DNA repair mechanisms and cellular function.

Main Methods:

• Generated a knockin mouse model with Nthl1 D227Y using CRISPR-cas9 genome editing.
• Utilized murine embryonic fibroblasts (MEFs) from heterozygous (Y/+) and homozygous (Y/Y) mutant mice.
• Assessed double-strand breaks, genomic instability, replication stress, and proliferation.

Main Results:

• Nthl1 D227Y heterozygous and homozygous MEFs showed increased double-strand breaks and genomic instability.
• Replication stress and impaired proliferation were observed in both mutant genotypes.
• The D227Y variant appears to interfere with wild-type NTHL1 repair activity, potentially by shielding lesions.

Conclusions:

• The NTHL1 D227Y variant leads to significant cellular defects, including DNA damage and impaired proliferation.
• Both heterozygous and homozygous carriers of the NTHL1 D239Y mutation may face an elevated risk of DNA damage-associated diseases, such as cancer.