engleski

GCNA (ACRC) is an essential protease that repairs DNA-protein crosslinks during zebrafish embryonic development

Proteins that interact with DNA can become irreversibly covalently bound to DNA and form lesions called DNA-protein crosslinks (DPCs). Several groups have characterized the proteins Wss1 in yeast and SPRTN in metazoans as the first identified proteases involved in DPC repair. Impaired DPC repair can lead to DNA double-strand breaks, genomic instability, and consequently cell death, as well as cancer, premature aging, and neurodegenerative diseases at the organismal level. Recent studies have shown that there is a second potential DPC protease in metazoans: GCNA (ACRC). Phylogenetic analysis and structural modeling have revealed similarities between SPRTN and GCNA. The aim of our study was to determine whether GCNA is involved in DPC repair in vivo and whether its putative protease core is important for GCNA function. We used zebrafish (Danio rerio) as a model organism, CRISPR/Cas9 gene manipulation to generate zebrafish mutant strains, and the RADAR (rapid approach to DNA adduct recovery) method to isolate DPCs from mutant embryos. We introduced a mutation in the putative protease active site of GCNA that includes a deletion of E451 (putative catalytic glutamate). Our results showed that the mutant embryos had 100% mortality within the first 24 hours of development due to maternal effect. This phenotype was rescued by mRNA encoding wild-type GCNA protein. Notably, the mutant embryos had an increased total amount of cellular DPCs. This study demonstrates the importance of GCNA for DPC repair at the organismal level during embryonic development and the importance of the putative protease core for GCNA function.

DNA repair ; DNA-protein crosslinks (DPCs) ; GCNA (ACRC) protease ; CRISPR/Cas9 ; zebrafish

nije evidentirano