Modulation of riboflavin biosynthesis and utilization in mycobacteria

Area of Science:

• Microbiology
• Biochemistry
• Immunology

Background:

• Riboflavin (vitamin B2) is vital for cellular redox metabolism, synthesized de novo by microbes like Mycobacterium tuberculosis (Mtb).
• Riboflavin pathway intermediates activate mucosal-associated invariant T (MAIT) cells, linking microbial metabolism to immune responses.

Approach:

• Conditional knockdowns (hypomorphs) of riboflavin biosynthesis and utilization genes were created in Mycobacterium smegmatis (Msm) and Mtb using inducible CRISPR interference.
• The study analyzed the impact of gene silencing on microbial viability, gene transcription, protein levels, and riboflavin production.

Key Points:

• Both Msm and Mtb can assimilate external riboflavin, even without a canonical transporter.
• Functional redundancy in lumazine synthase was observed in Msm.
• Silencing of ribA2 or ribF in Mtb proved highly bactericidal.
• In Msm, ribA2 silencing led to broad pathway gene knockdown, riboflavin depletion, and was also bactericidal.

Conclusions:

• The study validates riboflavin biosynthesis genes as potential drug targets for tuberculosis treatment.
• The generated hypomorph collection serves as a valuable tool for studying riboflavin pathway roles in mycobacterial physiology and MAIT cell activation.