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The effect of post translational modifications on WRN function and pathway choice in double strand break repair

Carlsbergfondets internationaliseringsstipendier

What

Werner Syndrome is a genetic disorder characterized by premature aging in adults and caused by a mutation in the WRN gene. WRN is associated with genomic instability and have been shown to be important for DNA repair. Double strand breaks (DSBs) are one of the major lesions that account for most of the deletions/insertions in the genome and chromosomal rearrangements. WRN is very important in the repair pathways of these lesions, however there are still many unanswered questions regarding its regulation and its effect on DSB repair pathway choice. In this project we aim to investigate how cell cycle induced post translational modifications (PTMs) regulate WRN and what effect it has on DSB repair.

Why

The growing proportion of elderly people represent an increasing economic challenge, not least because of age-associated diseases that poses a significant cost to society. Genetic age-accelerated diseases, such as Werner Syndrome, offer a great opportunity to study and understand the molecular mechanisms that lead to aging. This project will provide us with new insight on the mechanisms behind the regulation of WRN and help elucidate its important role in the DNA damage response network. The discoveries can result in the development of new treatments, not only for patients with accelerated aging, but also for age-related diseases.

How

Cell cycle induced phosphorylations will be identified using a combination of bioinformatics and mass spectroscopy. The effect of these on WRN function will be assessed by creating mutants that either mimic phosphorylation or are unable to be phosphorylated. These mutants will be used to determine the effect on WRN enzymatic activities (i.e., helicase, ATPase, ssDNA annealing, exonuclease) and the effect on recruitment to DBSs and on DSB repair pathways, such as non-homologous end joining and homologous recombination. Protein interaction partners will be identified using immunoprecipitation and mass spectroscopy. Other PTMs such as acetylations and ubiquitylations will also be identified and their functional effects on WRN and DSB repair will be evaluated.

SSR

Determining the underlying cellular and molecular processes that deteriorate with age and lead to increased disease susceptibility is critical to meet the growing healthcare needs of the aging human population.