Optimal construction of a functional interaction network from pooled library CRISPR fitness screens

Functional interaction networks, where edges connect genes likely to operate in the same biological process or pathway, can be inferred from CRISPR knockout screens in cancer cell lines. Genes with similar knockout fitness profiles across a sufficiently diverse set of cell line screens are likely to be co-functional, and these "coessentiality" networks are increasingly powerful predictors of gene function and biological modularity. While several such networks have been published, most use different algorithms for each step of the network construction process. In this study, we identify an optimal measure of functional interaction and test all combinations of options at each step - essentiality scoring, sample variance and covariance normalization, and similarity measurement - to identify best practices for generating a functional interaction network from CRISPR knockout data. We show that Bayes Factor and Ceres scores give the best results, that Ceres outperforms the newer Chronos scoring scheme, and that covariance normalization is a critical step in network construction. We further show that Pearson correlation, mathematically identical to ordinary least squares after covariance normalization, can be extended by using partial correlation to detect and amplify signals from "moonlighting" proteins which show context-dependent interaction with different partners.