Functional interrogation of DNA damage response variants with base editing screens

Area of Science:

• Genetics
• Molecular Biology
• Cancer Research

Background:

• DNA damage response (DDR) gene mutations compromise genome integrity, increasing cancer and genetic disorder risks.
• Understanding DDR gene function is crucial for disease research.

Purpose of the Study:

• To identify novel variants in DDR genes using CRISPR-based screening.
• To investigate the functional impact of these variants on cellular fitness and genome stability.
• To uncover new DDR gene functions and their roles in human diseases.

Main Methods:

• CRISPR-dependent cytosine base editing screens were employed.
• >2,000 sgRNAs were used to generate nucleotide variants in 86 DDR genes.
• Cellular fitness assays were performed upon DNA damage induction.

Main Results:

• Over 2,000 variants in 86 DDR genes were identified, affecting cellular fitness.
• Loss- and gain-of-function mutants in 53BP1's Tudor domain were discovered, impacting USP28 binding.
• TRAIP variants defining a domain conferring resistance to topoisomerase I inhibition were characterized.
• ATM kinase mutations with opposing phenotypes and CHK2 loss-of-function mutations were identified.

Conclusions:

• This study provides a valuable resource for discovering DDR gene functions.
• The identified variants offer insights into DDR gene roles in cancer and genetic disorders.
• Further research on these DDR variants can expedite studies in human disease.