Due to evolutionary conservation of biology, experimental knowledge captured from genetic studies in eukaryotic model organisms provides insight into human cellular pathways and ultimately physiology. Yeast chemogenomic profiling is a powerful approach for annotating cellular responses to small molecules. Using an optimized platform, we provide the relative sensitivities of the heterozygous and homozygous deletion collections for nearly 1800 biologically active compounds. The data quality enables unique insights into pathways that are sensitive and resistant to a given perturbation, as demonstrated with both known and novel compounds. We present examples of novel compounds that inhibit the therapeutically relevant fatty acid synthase and desaturase (Ole1p and Fas1p), and demonstrate how the individual profiles facilitate hypothesis-driven experiments to delineate compound mechanism of action. Importantly, the scale and diversity of tested compounds yields a dataset where the number of modulated pathways approaches saturation. This resource can be used to map novel biological connections, and also identify functions for unannotated genes. We validated hypotheses generated by global 2-way hierarchical clustering of profiles for (i) novel compounds with a similar mechanism of action acting upon microtubules or vacuolar ATPases, and (ii) an un-annotated ORF, YIL060w, that plays a role in respiration in the mitochondria. Finally, we identify and characterize background mutations in the widely used yeast deletion collection which should improve the interpretation of past and future screens throughout the community. This comprehensive resource of cellular responses enables the expansion of our understanding of eukaryotic pathway biology.

Table S1

Compounds used in this study

Table_S1.xls

Table_S2

Hit frequencies of HIP and HOP strains

Table_S3

v-ATPase inhibitor correlation list

Table_S4

microtubule inhibitor compounds as identified by HIP HOP

Table_S5

Summary of identified HIP strains with common background mutations

Table_S6

Signature hits for inhibition of non-proteinaceous targets

HIP_scores-benomyl

Tab delimited text file with the data of all performed Benomyl experiments

HOP_scores-benomyl

Tab delimited text file with the data of all performed Benomyl experiments

HIP_scores

Tab-delimited text file with MADL and z-scores for all approx. 1800 compounds tested in this study

HOP_scores

Tab-delimited text file with MADL and z-scores for all approx. 1800 compounds tested in this study

HIPHOP_gene_correlation

Delimited text file depicting calculated correlation of genes in relation to treatment with the 1800 compounds tested in this study

Changes after Nov 19, 2014: The web version of the HIP HOP datasets had to be taken offline because it was no longer compatible with current web safety standards. The gene-gene correlation table, which had previously been accessible only through that web version, has now been uploaded to ensure continued access for the scientific community. For table structure and how it was calculated consult the provided “HipHop_gene_correlation_README.docx” and "HipHop_gene_correlation_README.pdf" files.