UCSF

Bacterial essential genes contribute to the most fundamental processes of cellular life. The study of their functions in vivo has long been intractable to systematic genetic approaches, which are fundamental to understanding pathway level connections that govern cellular life and are a requirement for dissecting the complex cellular processes to which essential genes contribute. In Chapter 1 of this work I review recent advances in mapping gene-phenotype relationships in bacteria using the CRISPR-based technology, CRISPR interference (CRISPRi) for titratable gene knockdowns, focusing on their applications to the studies of essential genes, the exploration of chemical-genetic interactions, and the prospects for disentangling complex phenotypes in diverse bacterial species. In Chapter 2 I describe my analysis of the essential gene functions in the model Gram-negative bacterium Escherichia coli and the model Gram-positive Bacillus subtilis using datasets from paired chemical-genetic screens. In this work I identify both shared and Gram-negative specific mechanisms of collateral sensitization to antibiotic action. In Chapter 3 I investigate a fundamental property of essential genes, which is the relationship between their expression level and the cellular growth rate. Here, further developing CRISPRi tools in bacteria to predictably titrate knockdown efficacy, I interpret the knockdown-fitness relationships of each essential gene in E. coli and B. subtilis, discovering broad conservation of constraints setting and maintaining expression levels across these diverged species.