This variability highlights the benefits of aggregating information across cell lines. Although, six well-known cell cycle blockers were detected in all cell lines delineating a module core in CODIM1, 26 additional cell cycle blockers were only detected as part of this module in one or two of the three cell lines (shaded columns). Grey: compounds with a known effect on cell cycle (Supplementary Table 5). (A) Heatmap: expression fold change in each cell line with shaded columns, indicating chemical treatments not present in the respective cell-line-specific module.
Prediction of novel cell cycle inhibitors based on CODIM1. Total intensity of fluorescent signal in segmented spots/area has been quantified as indicated with arrows. Cells have been transfected for 48 h with siRNA targeting indicated gene, and experiments of (C) LDL-uptake assay with fluorescently labeled DiI–LDL and (D) staining with cholesterol binding dye filipin was performed. (C, D) Representative image of fixed cells taken by automated widefield fluorescence microscopy.
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Genes subjected to functional knockdown assays of LDL uptake and free cellular cholesterol levels are highlighted in red or orange depending on whether validated as a functional regulator or not, respectively (two or more siRNAs having consistent effect with absolute z-score >1 and FDR-corrected P-value <0.01, Supplementary Table 6). The second component (enriched in green set) is enriched in ER-related processes, such as protein N-linked glycosylation, transport and unfolded protein response. (B) Projection onto the STRING protein–protein network reveals a densely interconnected component (enriched in blue set) corresponding to fatty acid and (chole-)sterol metabolism (Supplementary Table 5 and Supplementary ). Hierarchical clustering of genes from CODIM2 shows two major components (labeled in blue and green) and three smaller groups (dark green, brown, light blue).
Black boxes indicate gene membership of cell-line-specific modules. (A) Heatmap of drug-induced gene expression changes (fold change) of CODIM2. Novel functional roles of uncharacterized genes as functional regulators of cellular cholesterol levels. We predicted and experimentally validated novel cell cycle inhibitors and modulators of PPARγ, estrogen and adrenergic receptors, with potential for developing new therapies against diabetes and cancer. A conserved module associated with (chole)sterol metabolism revealed novel regulators of cellular cholesterol homeostasis 10 of them were validated in functional imaging assays.Īnalysis of drugs clustered into modules can give new insights into their mechanisms of action and provide leads for drug repositioning. About 70% of modules are conserved across cell lines, a lower bound of 15% was estimated for their conservation across organisms, and between the in vitro and in vivo systems.ĭrug-induced transcriptional modules can predict novel gene functions. Identifying drug-induced transcriptional modules separately in three human cell lines and rat liver allows assessment of their conservation across model systems. Biclustering of drug-induced gene expression profiles resulted in modules of drugs and genes, which were enriched in both drug and gene annotations.
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