Metabolism is a fundamental cellular mechanism. Its regulation is crucial to maintain cellular homeostasis in response to fluctuation in energy demands and in the availability of nutrients. However, metabolic reactions can be harmful for cells, for example by leading to an increase in oxidative stress or through the generation of toxic by-products, which can in turn damage DNA. To deal with these insults, cells have evolved sophisticated DNA damage response (DDR) pathways that allow for the maintenance of genome integrity. Recent years have seen remarkable progress in unraveling the diverse mechanisms of the DDR. Through such work, it has also emerged that cellular metabolic regulation not only generates DNA damage but also impacts on DNA repair, yet a systematic analysis aimed at identifying interactions between metabolic alterations and the DDR remains unreported. Here, we have functionally explored such interplay, by taking a global and unbiased genetic approach.

By utilizing a pooled CRISPR-Cas9 sgRNA library targeting some 3,000 metabolic genes, we have determined the effects on DNA repair following the induction of DNA double-strand breaks induced by the chemotherapeutic etoposide, an inhibitor of topoisomerase 2. Through a phenotypic FACS-based screen with gH2AX used as a marker of DNA damage, we have identified metabolic genes that are required for DNA repair. Candidate genes have been prioritized and validated via an arrayed CRISPR screen based on high-throughput microscopy measuring multiple parameters, across different cell lines and DNA double-strand break inducing agents. We will present data on how metabolic genes impact on DNA repair.

Our research sheds light on how the fundamental cellular process of DNA damage maintenance is affected by alterations in cellular metabolism. Moreover, understanding the dynamic interplay between metabolic factors and DDR is also of crucial importance to better design efficient cancer treatments, since cancers are both genetic and metabolic diseases.