Figure 7. Comparison of DUB knockout and inhibition.

(A) The number of significantly differentially expressed genes (adjusted p-value <0.05) as a result of small molecule DUB inhibition (24 hr treatment) and knockout of the putative target (96 hours after transfection with guide). (B) Hierarchical clustering of log2FC values for significantly differentially expressed genes (adjusted p-value <0.05) for small molecule inhibitors of DUBs, the knockout of the putative DUB targets of the small molecules, and the DUB knockout hits that resulted in more than 20 differentially expressed genes. inhibition with XL177A, the DUB inhibitor and knockout were strongly correlated, but the inhibitor resulted in substantially more DE genes, and (iii) in five cases (inhibition of COPS5 with Compound 6 or CSN5i-3, inhibition of OTUD7B with I-145, inhibition of USP19 with I-124, and inhibition of USP30 with MF-094) the inhibitor and knockout had dissimilar signatures (a and b). CRISPR-Cas9 mediated knockout or small molecule inhibition of USP14 with I-335 resulted in only two DE genes, and the most strongly perturbed DE gene was the same in both instances: UBC. From these data, we conclude that I-335 is a potent and selective inhibitor of USP14.

Figure 7—figure supplement 1. Gene set enrichment results (Hypergeometric test) of the top seven co-dependent genes for all DUBs using GO Molecular Function gene sets.