Bioactive compounds are widely used to modulate protein function and can serve as important leads for drug development. Identifying the in vivo targets of these compounds remains a challenge. Using yeast, we integrated three genome-wide gene-dosage assays to measure the effect of small molecules in vivo. A single TAG microarray was used to resolve the fitness of strains derived from pools of (i) homozygous deletion mutants, (ii) heterozygous deletion mutants and (iii) genomic library transformants. We demonstrated, with eight diverse reference compounds, that integration of these three chemogenomic profiles improves the sensitivity and specificity of small-molecule target identification. We further dissected the mechanism of action of two protein phosphatase inhibitors and in the process developed a framework for the rational design of multidrug combinations to sensitize cells with specific genotypes more effectively. Finally, we applied this platform to 188 novel synthetic chemical compounds and identified both potential targets and structure-activity relationships.