Populations of isogenic cells often respond coherently to signals despite differences in protein abundance and cell state. Our previous work in the Saccharomyces cerevisiae pheromone response system (PRS) uncovered processes that reduced cell-to-cell variation in signal and response. To understand these and other processes that controlled variation, we generated a whole-genome collection of haploid strains with deletions in non-essential genes and used high-throughput flow cytometry to screen more than 1000. We identified 50 'variation genes' required for normal cell-to-cell variability in signal and response. Some genes affected only signal variability, signal strength, or system output, defining these quantities as separable 'axes' of system behavior. Two genes affected cytoplasmic microtubule function.