Abstract: We demonstrate the efficacy of a genome-wide protocol in yeast that allows the identification of those gene products that functionally interact with small molecules and result in the inhibition of cellular proliferation. Here we present results from screening 10 diverse compounds in 80 genome-wide experiments against the complete collection of heterozygous yeast deletion strains. These compounds include anticancer and antifungal agents, statins, alverine citrate, and dyclonine. In several cases, we identified previously known interactions; furthermore, in each case, our analysis revealed novel cellular interactions, even when the relationship between a compound and its cellular target had been well established. In addition, we identified a chemical core structure shared among three therapeutically distinct compounds that inhibit the ERG24 heterozygous deletion strain, demonstrating that cells may respond similarly to compounds of related structure. The ability to identify on-and-off target effects in vivo is fundamental to understanding the cellular response to small-molecule perturbants.