Understanding the Heterogeneity in Gene Regulatory Responses to Misfolded Protein Toxicity

Protein misfolding is a problem across all organisms, but the reasons behind misfolded protein (MP) toxicity to cells are largely unknown. To better understand toxicity, I investigate if toxicity from MPs affects all cells equally or affects some cell subpopulations more than others, such as older cells. To define cell subpopulations, I optimized a cutting-edge single-cell RNA sequencing platform (scRNAseq) for yeast. By using scRNAseq in yeast, I studied the expression variability of many genes across populations of thousands of cells. I studied how the transcriptomes of single cells differ from one another in various conditions: at different stages in the growth phase and with different engineered MPs. Differences in gene expression between strains expressing misfolded vs. properly folded proteins were found, confirming previous proteomic data. Further, I found a greater number of cell subpopulations in a MP expressing strain compared to a properly folded protein expressing strain, implying more differentiated subpopulations, potentially in response to toxicity from MPs. This observation is consistent with previous observations that heterogeneity within microbial populations can be beneficial to their fitness by allowing that population to thrive in stressful environments. Thus, my data provide insights about evolutionary biology and how strains respond to stress. Further, after identifying subpopulations with a more severe transcriptional response to MPs, I studied the cells’ physiology to gain insights about why that subpopulation is sensitive to MPs and found an upregulation of markers of aging, stress response, and shortening of lifespan. Observing characteristics of cell subpopulations, I also found differences dependent on stages of the cell cycle. Overall, this study provides insights on the gene regulatory responses associated with MP toxicity by revealing which type of cells are most sensitive to this intracellular threat.