Abstract

A central remaining question in the post-genomic era is how genes interact to form biological pathways. Measurements of gene dependency across hundreds of cell lines have been used to cluster genes into co-essential pathways, but this approach has been limited by ubiquitous false positives. Here, we develop a statistical method that enables robust identification of gene co-essentiality and yields a genome-wide set of functional modules. This almanac recapitulates diverse pathways and protein complexes and predicts the functions of 102 uncharacterized genes. Validating top predictions, we show that TMEM189 encodes plasmanylethanolamine desaturase, the long-sought key enzyme for plasmalogen synthesis. We also show that C15orf57 binds the AP2 complex, localizes to clathrin-coated pits, and enables efficient transferrin uptake. Finally, we provide an interactive web tool for the community to explore the results (coessentiality.net). Our results establish co-essentiality profiling as a powerful resource for biological pathway identification and discovery of novel gene functions.