Genome-wide gene expression tuning reveals diverse vulnerabilities of M. tuberculosis

Summary

Antibacterial agents target the products of essential genes but rarely achieve complete target inhibition. Thus, the all-or-none definition of essentiality afforded by traditional genetic approaches fails to discern the most attractive bacterial targets: those whose incomplete inhibition results in major fitness costs. In contrast, gene “vulnerability” is a continuous, quantifiable trait that relates the magnitude of gene inhibition to the effect on bacterial fitness. We developed a CRISPR interference-based functional genomics method to systematically titrate gene expression in Mycobacterium tuberculosis (Mtb) and monitor fitness outcomes. We identified highly vulnerable genes in various processes, including novel targets unexplored for drug discovery. Equally important, we identified invulnerable essential genes, potentially explaining failed drug discovery efforts. Comparison of vulnerability between the reference and a hypervirulent Mtb isolate revealed incomplete conservation of vulnerability and that differential vulnerability can predict differential antibacterial susceptibility. Our results quantitatively redefine essential bacterial processes and identify high-value targets for drug development.

Graphical abstract

Highlights

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  Titratable CRISPRi enables quantification of target vulnerability in mycobacteria

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  Essential genes and processes vary widely in their vulnerability

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  Differential vulnerability predicts differential antibacterial susceptibility

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  Generalizable approach allows prioritization of high-value targets for drug discovery

Abstract

Conventional genetic approaches rely on complete gene inactivation to identify potential antimicrobial targets. In contrast to this all-or-nothing effect, small molecules rarely achieve complete target inhibition. Here, a CRISPR interference system using M. tuberculosis as a model organism is used to titrate gene expression and uncover gene vulnerability, redefining the concept of essential genes and identifying antimicrobial targets.