Year: 2026
Type: preprint
Abstract: ABSTRACT Mycobacterium tuberculosis has a flavin-intensive metabolism, with more than 180 proteins that require flavin or deazaflavin cofactors. M. tuberculosis requires riboflavin biosynthesis for growth in standard culture conditions in vitro , which can be overcome by providing exogenous riboflavin. It is unknown whether M. tuberculosis also requires riboflavin synthesis to grow or persist in infected host cells or tissues. Here we used a M. tuberculosis strain that conditionally expresses ribA2 , which encodes a bifunctional enzyme that catalyzes the first steps in both branches of riboflavin biosynthesis, to determine whether M. tuberculosis requires riboflavin biosynthesis during infection. Transcriptional repression of ribA2 caused death of M. tuberculosis in vitro without riboflavin supplementation. RNAseq revealed that riboflavin starved M. tuberculosis induce a transcriptional program indicating decreased electron transport chain function and broad changes to central metabolism. Transcriptional repression of ribA2 prevented M. tuberculosis growth in cultured macrophages and caused clearance of M. tuberculosis from infected tissues at all stages of infection in both C57BL/6 mice that form cellular lung lesions and C3HeB/FeJ mice that develop necrotic lung granulomas. Our results demonstrate that M. tuberculosis requires RibA2 for viability at all stages of infection and support further characterization of the riboflavin biosynthesis pathway as a potential target for development of new anti-tubercular drugs. AUTHOR SUMMARY Tuberculosis (TB) caused by Mycobacterium tuberculosis kills over 1.2 million people worldwide each year, making it the deadliest bacterial infection. TB treatment involves combinations of antibiotics taken for at least six months. M. tuberculosis readily becomes resistant to antibiotics by mutation and multidrug resistant strains account for nearly 4% of new TB cases. New antibiotics for TB are needed to combat drug resistance and shorten TB treatment. Riboflavin biosynthesis is an attractive target for antibiotic development because mammals do not synthesize riboflavin, instead obtaining it from the diet. Here, using a M. tuberculosis strain that conditionally expresses the riboflavin biosynthesis enzyme RibA2, we show that inhibition of riboflavin biosynthesis kills M. tuberculosis in lab culture conditions and in mice at all stages of infection. Our results demonstrate that riboflavin starvation is bactericidal for M. tuberculosis and identify the riboflavin biosynthesis pathway as a promising target for new drugs to treat TB.
Institution: University of Minnesota
Language: English
Subfield: Infectious Diseases
Field: Medicine
Domain: Health Sciences
Open Access status: green
OpenAlex ID: W7214477629