Systematic morphological profiling of human gene and allele function via Cell Painting

• Published in: eLife 2017-03, Vol.6
• Main Authors: Rohban, Mohammad Hossein, Singh, Shantanu, Wu, Xiaoyun, Berthet, Julia B, Bray, Mark-Anthony, Shrestha, Yashaswi, Varelas, Xaralabos, Boehm, Jesse S, Carpenter, Anne E
• Format: Article
• Language: English
• Subjects: Adaptor Proteins, Signal Transducing - genetics; Adaptor Proteins, Signal Transducing - metabolism; Alleles; Annotations; Cell Line, Tumor; Cell Nucleus - metabolism; Cell Nucleus - ultrastructure; Cell Transformation, Neoplastic - genetics; Cell Transformation, Neoplastic - metabolism; Cell Transformation, Neoplastic - pathology; Cloning; Computational and Systems Biology; Disease; DNA sequencing; Endoplasmic Reticulum - metabolism; Endoplasmic Reticulum - ultrastructure; Fluorescent Dyes - chemistry; Gene clusters; Gene expression; Gene Expression Profiling; Gene Expression Regulation; gene functional annotation; Genomes; Golgi Apparatus - metabolism; Golgi Apparatus - ultrastructure; HEK293 Cells; Hippo Pathway; Humans; Image Processing, Computer-Assisted; Intracellular Signaling Peptides and Proteins - genetics; Intracellular Signaling Peptides and Proteins - metabolism; Methods; Mitochondria - metabolism; Mitochondria - ultrastructure; morphological profiling; Morphology; Multigene Family; NF-kappa B - genetics; NF-kappa B - metabolism; NF-kB pathway; NF-κB protein; Observations; Optical Imaging; Osteoblasts - metabolism; Osteoblasts - pathology; Phosphoproteins - genetics; Phosphoproteins - metabolism; Physiological aspects; Protein-Serine-Threonine Kinases - genetics; Protein-Serine-Threonine Kinases - metabolism; Proto-Oncogene Proteins c-rel - genetics; Proto-Oncogene Proteins c-rel - metabolism; Raf protein; Signal Transduction; Staining and Labeling - methods; TNF Receptor-Associated Factor 2 - genetics; TNF Receptor-Associated Factor 2 - metabolism; Tools and Resources; TRAF2 protein; Trans-Activators; Transcription; Transcription Factors; Yes-associated protein
• ISSN: 2050-084X; 2050-084X
• DOI: 10.7554/elife.24060

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.