Dissecting the yeast transcriptome landscape through genetic and environmental Perturb-seq analyses

Cellular heterogeneity is a key determinant for disease outcome and therapeutic treatments across virtually all organisms. Variability within a population underlies the different cellular responses that ultimately determine cell fate and phenotypic diversity. Little is known about the molecular basis of such cell-to-cell variability nor how it impacts the phenotypic spectrum that emerges during adaptive responses to environmental fluctuations. To understand the contribution of each gene to the resulting adaptive transcriptome, we designed a gene-deletion strategy to combine genetic x environmental perturbation screens with single cell RNA-seq profiling (scRNA-seq). Here we profiled a total of 1.2M cells from more than 3000 different genotypes under normal and osmostress conditions to generate high resolution genotype-transcriptome maps. We use transcriptional phenotype to identify distinct transcriptional architecture, variable gene usage, gene function associations, and uncover regulators of heterogeneity. Our results demonstrate that the majority of core osmoresponsive genes are only used by a fraction of the population, including the restricted use of transcription factor families. We leveraged inter genotype transcriptome correlation to predictions of gene function. Harnessing intra- genotype heterogeneity led us to uncover and experimentally validate both positive and negative universal or condition-specific regulators of cellular heterogeneity. Our findings expose the complexity of gene and genotype transcriptome layers.