However, over 30% of genes in the human body do not have a known role. Genome sequencing can determine the exact order of these bases, and has revealed a long list of mutations in genes that could cause particular diseases. Humans have about 30,000 genes built from DNA, which contain specific sequences of bases. This information is stored as a code made up of four chemicals called bases. A genome is the complete DNA set of an organism, which contains all the information to build the body and keep it working. Many human diseases are caused by particular changes, called mutations, in patients’ DNA. We make the images and raw data publicly available, providing an initial morphological map of major biological pathways for future study. 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. This unbiased morphologic map of gene function revealed TRAF2/c-REL negative regulation of YAP1/WWTR1-responsive pathways. We used novel subpopulation-based visualization methods to interpret the morphological changes for specific clusters. 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 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. (H): All genes/alleles in Cluster 8 and 10 induce cell rounding. (I) The NF-κB signaling pathway is the most enriched when searching for gene overexpressions that downregulate known YAP/TAZ targets (CYR61, CTGF, and BIRC5). Genes shown here are only those where either the wild-type gene or its constitutively activating allele yielded a phenotype distinct from controls. (C) Pathways sorted based on proportion of their associated gene showing a detectable phenotype. (D) Highly correlated proteins (according to morphology in the Cell Painting assay) that have also been reported to interact physically. (E) Highly correlated genes (according to morphology in the Cell Painting assay) that have also been annotated to be related to the same pathway. (F) Gene Ontology terms associated with each gene cluster ( Alexa and Rahnenführer, 2009). (G) Rank ordered list of distinctive features based on their z-scores for Cluster 19. Constitutively active mutant annotations were obtained by literature search for all the mutants in the experiment showing a detectable phenotype. (A) List of all the 323 constructs used in the experiment along with the target transcript and their public clone ID. (B) Replicate correlation is higher in the constitutively active mutant allele compared to the wild-type allele, except for AKT3_E17K. Supplementary file 1: Supporting and supplemental data for the figures and experiments.
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