Investigating the Conditionally Essential Role of Human METAP1

Abstract

CRISPR‐based forward genetic screens make it possible to systematically identify genes required for survival and growth in human cells, and therefore provide a powerful approach to characterize protein function and uncover targetable vulnerabilities in human cancer cells. Notably, it has become appreciated that gene essentiality can be context‐dependent, as CRISPR‐based loss‐of‐function screens across hundreds of human cancer cell lines have uncovered cell‐essential genes that can vary with cell‐intrinsic characteristics such as genotype and lineage. Importantly, environmental factors also influence cell physiology. However, there has been little consideration into how nutrient availability might impact gene essentiality, and most CRISPR‐based screens of human cells are performed in vitro using culture media that poorly recapitulate nutrient conditions in the body. To test the hypothesis that medium composition influences gene essentiality, our lab recently performed genome‐wide CRISPR screens of human blood cancer cell lines in traditional versus Human Plasma‐Like Medium (HPLM), a physiologic medium that we previously developed to more closely reflect the metabolic composition of human blood. Among the strongest HPLM‐essential hits identified from our screen results is METAP1, which encodes one of two human enzymes (METAP1 and METAP2) that can catalyze the cleavage of N‐terminal methionine from nascent polypeptides. Interestingly, these two METAPs are co‐expressed across most human cell lines, are each localized to the cytosol, and also share seemingly similar substrate preferences. However, our screen results indicate that endogenous METAP2 cannot complement the loss of METAP1in HPLM‐cultured cells, suggesting that METAP1 serves a non‐redundant and context‐dependent cell‐essential role. Therefore, we hypothesize that identifying the gene‐nutrient interaction that underlies conditional METAP1essentiality will provide key insights into this unforeseen role. Importantly, we have engineered METAP1‐knockout cells and, in turn, confirmed the conditional CRISPR phenotype for METAP1by performing relative growth assays. Therefore, we are now poised to identify the gene‐nutrient interaction and will proceed to use tools in biochemistry, metabolomics, and molecular biology to explore the context‐dependent role of METAP1 in proliferating human cells.