The complex human ubiquitin proteasome system, comprised of over 700 ubiquitin ligases (E3s) and deubiquitinases (DUBs), has been difficult to systematically and phenotypically characterize. We performed a chemical-genetic CRISPR-Cas9 screen to identify E3s/DUBs whose loss renders cells sensitive or resistant to 41 compounds targeting a broad range of biology, including cytoskeletal integrity, mitosis, G1/S progression, genome stability, translation, metabolism, and vesicular transport.  Genes and compounds both clustered functionally, showing that inhibitors of related pathways interact similarly with the E3s/DUBs and demonstrating the robustness of our screen. Some genes, such as FBXW7, showed interactions with more than one-third of the compounds. Others, such as the mostly un-studied RNF25 and FBXO42, showed interactions primarily with a single compound (MMS for RNF25) or a set of related compounds (mitotic cluster for FBXO42). Mutation of several E3s with sensitivity to mitotic inhibitors had an increase in aberrant mitoses, suggesting a role for these genes in cell cycle regulation. Overall, our comprehensive CRISPR-Cas9 screen uncovered 466 gene-compound interactions covering 25% of the E3s/DUBs interrogated.