Mapping the Cellular Response to Small Molecules Using Chemogenomic Fitness Signatures

Abstract

Genome-wide characterization of the in vivo cellular response to perturbation is fundamental to understanding how cells survive stress. Identifying the proteins and pathways perturbed by small molecules affects biology and medicine by revealing the mechanisms of drug action. We used a yeast chemogenomics platform that quantifies the requirement for each gene for resistance to a compound in vivo to profile 3250 small molecules in a systematic and unbiased manner. We identified 317 compounds that specifically perturb the function of 121 genes and characterized the mechanism of specific compounds. Global analysis revealed that the cellular response to small molecules is limited and described by a network of 45 major chemogenomic signatures. Our results provide a resource for the discovery of functional interactions among genes, chemicals, and biological processes.

Structural elements that govern Sec14-like PITP sensitivities to potent small molecule inhibitors

“…Briefl y, Escherichia coli BL21 (DE3) cells carrying the appropriate (PtdCho) in vitro, and it coordinates PtdIns-4-phosphate production with PtdCho metabolism so as to promote membrane traffi cking through trans-Golgi network and endosomal compartments in vivo (17)(18)(19). Of the SMIs that target Sec14 activity, the nitrophenyl(4-(2-methoxyphenyl) piperazin-1-yl)methanones (NPPMs) are best understood ( 14,20,21 ). These SMIs inhibit Sec14 by loading into the phospholipid binding pocket of this PITP and preventing subsequent rounds of lipid exchange ( 14 ).…”

Beyond Agar: Gel Substrates with Improved Optical Clarity and Drug Efficiency and Reduced Autofluorescence for Microbial Growth Experiments

“…Microbial cultures can be maintained in liquid growth medium or on solid/semisolid gel substrates (3). Liquid colony maintenance allows the microbial cultures to expand rapidly and is used in screens that rely on optical density measurements to extract kinetic growth parameters or use downstream liquid-based assays such as flow cytometry or high-throughput microscopy (4)(5)(6)(7). While throughput in liquid is generally bound to a maximum of 96 or 384 samples per plate, solid-substrate colony maintenance has the advantage that individual cell clones can be readily separated and that colony parameters such as color, shape, structure, and size can be extracted easily using digital image acquisition (8)(9)(10).…”

Off-Target Effects of the Septin Drug Forchlorfenuron on Nonplant Eukaryotes

“…Alternatively, septin-targeting molecules may be among the compounds produced naturally by plants to resist septin-facilitated infection by certain fungi (67) or by pathogenic bacteria to counteract septin-based host defense mechanisms in metazoans (68). Finally, a very recent chemical genomics screen identified a synthetic small molecule that inhibited the growth of diploid budding yeast cells in which one copy of the septin-encoding gene CDC12 was deleted (69). This compound, 1-ethyl-3-(4-methoxyphenyl)-6-methylpyrimido [5,4-e][1,2,4]triazine-5,7-dione, which in many ways resembles guanosine, was found to be ϳ5-fold more potent than FCF in inhibiting human septin function in a wound-healing assay using cultured cells (69).…”

“…Finally, a very recent chemical genomics screen identified a synthetic small molecule that inhibited the growth of diploid budding yeast cells in which one copy of the septin-encoding gene CDC12 was deleted (69). This compound, 1-ethyl-3-(4-methoxyphenyl)-6-methylpyrimido [5,4-e][1,2,4]triazine-5,7-dione, which in many ways resembles guanosine, was found to be ϳ5-fold more potent than FCF in inhibiting human septin function in a wound-healing assay using cultured cells (69). Despite this improved efficacy, however, the overall cellular response signature generated by this newly identified septin drug, i.e., the targeted biological process that emerges when considering all of the other yeast mutants that also were hypersensitive to the chemical, is one of iron homeostasis (69), in which septins have not been previously implicated.…”

“…This compound, 1-ethyl-3-(4-methoxyphenyl)-6-methylpyrimido [5,4-e][1,2,4]triazine-5,7-dione, which in many ways resembles guanosine, was found to be ϳ5-fold more potent than FCF in inhibiting human septin function in a wound-healing assay using cultured cells (69). Despite this improved efficacy, however, the overall cellular response signature generated by this newly identified septin drug, i.e., the targeted biological process that emerges when considering all of the other yeast mutants that also were hypersensitive to the chemical, is one of iron homeostasis (69), in which septins have not been previously implicated. Thus, the search continues for a small-molecule septin inhibitor lacking significant off-target effects.…”