Systematic morphological profiling of human gene and allele function via Cell Painting
- Broad Institute, United States
- Boston University School of Medicine, United States
- Novartis Institutes for BioMedical Research, United States
Abstract
We hypothesized that human genes and disease-associated alleles might be systematically functionally annotated using morphological profiling of cDNA constructs, via a microscopy-based Cell Painting assay. Indeed, 50% of the 220 tested genes yielded detectable morphological profiles, which grouped into biologically meaningful gene clusters consistent with known functional annotation (e.g., the RAS-RAF-MEK-ERK cascade). We used novel subpopulation-based visualization methods to interpret the morphological changes for specific clusters. This unbiased morphologic map of gene function revealed TRAF2/c-REL negative regulation of YAP1/WWTR1-responsive pathways. We confirmed this discovery of functional connectivity between the NF-κB pathway and Hippo pathway effectors at the transcriptional level, thereby expanding knowledge of these two signaling pathways that critically regulate tumor initiation and progression. We make the images and raw data publicly available, providing an initial morphological map of major biological pathways for future study.
Data availability
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Target Accelerator OEPublicly available at the Image Data Resource (http://idr-demo.openmicroscopy.org/about/).
Article and author information
Author details
Mohammad Hossein Rohban
Funding
National Science Foundation (NSF CAREER DBI 1148823)
- Anne E Carpenter
Broad Institute
- Anne E Carpenter
Carlos Slim Foundation
- Anne E Carpenter
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2017, Rohban et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Systematic morphological profiling of human gene and allele function via Cell Painting
eLife 6:e24060.
https://doi.org/10.7554/eLife.24060
Further reading
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Enhancers and promoters are classically considered to be bound by a small set of transcription factors (TFs) in a sequence-specific manner. This assumption has come under increasing skepticism as the datasets of ChIP-seq assays of TFs have expanded. In particular, high-occupancy target (HOT) loci attract hundreds of TFs with often no detectable correlation between ChIP-seq peaks and DNA-binding motif presence. Here, we used a set of 1003 TF ChIP-seq datasets (HepG2, K562, H1) to analyze the patterns of ChIP-seq peak co-occurrence in combination with functional genomics datasets. We identified 43,891 HOT loci forming at the promoter (53%) and enhancer (47%) regions. HOT promoters regulate housekeeping genes, whereas HOT enhancers are involved in tissue-specific process regulation. HOT loci form the foundation of human super-enhancers and evolve under strong negative selection, with some of these loci being located in ultraconserved regions. Sequence-based classification analysis of HOT loci suggested that their formation is driven by the sequence features, and the density of mapped ChIP-seq peaks across TF-bound loci correlates with sequence features and the expression level of flanking genes. Based on the affinities to bind to promoters and enhancers we detected five distinct clusters of TFs that form the core of the HOT loci. We report an abundance of HOT loci in the human genome and a commitment of 51% of all TF ChIP-seq binding events to HOT locus formation thus challenging the classical model of enhancer activity and propose a model of HOT locus formation based on the existence of large transcriptional condensates.
