CCAR1 promotes DNA repair via alternative splicing.

Area of Science:

• Molecular Biology
• Genetics
• Cell Biology

Background:

• DNA repair mechanisms are crucial for maintaining genomic stability.
• Homology-directed repair (HDR) and Fanconi anemia (FA) pathway are key DNA repair systems.
• CCAR1 is a poorly characterized gene with unknown functions in DNA repair.

Purpose of the Study:

• To identify novel factors indirectly regulating homology-directed repair (HDR).
• To investigate the role of CCAR1 in DNA repair and its underlying mechanisms.

Main Methods:

• Large-scale CRISPR inhibition and Cas9-editing screening of ~18,000 gene knockdowns.
• Analysis of CCAR1 depletion effects on HDR and interstrand crosslink repair.
• Western blot analysis of FANCA protein and mRNA levels.
• Transcriptomic analysis to assess global alternative splicing changes.

Main Results:

• CCAR1 depletion significantly reduced both HDR and interstrand crosslink repair.
• CCAR1 loss phenocopied the loss of the Fanconi anemia pathway, abrogating FANCA protein without affecting its mRNA.
• CCAR1 was found to prevent the inclusion of a poison exon in FANCA.
• CCAR1 regulates widespread alternative splicing changes that could damage coding sequences in human and mouse cells.

Conclusions:

• CCAR1 functions as a critical splicing fidelity factor.
• CCAR1 plays an essential, previously unrecognized role in maintaining genomic integrity through proper splicing regulation.