Redox-Dependent Condensation Of the Mycobacterial Nucleoid By WhiB4

Oxidative stress response in bacteria is generally mediated through coordination between the regulators of oxidant-remediation systems (e.g.OxyR, SoxR) and nucleoid condensation (e.g.Dps, Fis). However, these genetic factors are either absent or rendered nonfunctional in the human pathogenMycobacterium tuberculosis(Mtb). Therefore, howMtborganizes genome architecture and regulates gene expression to counterbalance oxidative imbalance during infection is not known. Here, we report that an intracellular redox-sensor, WhiB4, dynamically links genome condensation and oxidative stress response inMtb. Disruption of WhiB4 affects the expression of genes involved in maintaining redox homeostasis, central carbon metabolism (CCM), respiration, cell wall biogenesis, DNA repair and protein quality control under oxidative stress. Notably, disulfide-linked oligomerization of WhiB4 in response to oxidative stress activates the protein’s ability to condense DNAin vitroandin vivo. Further, overexpression of WhiB4 led to hypercondensation of nucleoids, redox imbalance and increased susceptibility to oxidative stress, whereas WhiB4 disruption reversed this effect. In accordance with the findingsin vitro, ChIP-Seq data demonstrated non-specific binding of WhiB4 to GC-rich regions of theMtbgenome. Lastly, data indicate that WhiB4 deletion affected the expression of only a fraction of genes preferentially bound by the protein, suggesting its indirect effect on gene expression. We propose that WhiB4 is a novel redox-dependent nucleoid condensing protein that structurally couplesMtb’sresponse to oxidative stress with genome organization and transcription.Significance StatementMycobacterium tuberculosis (Mtb)needs to adapt in response to oxidative stress encountered inside human phagocytes. In other bacteria, condensation state of nucleoids modulates gene expression to coordinate oxidative stress response. However, this relation remains elusive inMtb. We performed molecular dissection of a mechanism controlled by an intracellular redox sensor, WhiB4, in organizing both chromosomal structure and selective expression of adaptive traits to counter oxidative stress inMtb. Using high-resolution sequencing, transcriptomics, imaging, and redox biosensor, we describe how WhiB4 modulates nucleoid condensation, global gene expression, and redox-homeostasis. WhiB4 over-expression hypercondensed nucleoids and perturbed redox homeostasis whereas WhiB4 disruption had an opposite effect. Our study discovered an empirical role for WhiB4 in integrating redox signals with nucleoid condensation inMtb.