Please Note: The main page lists projects via BBSRC Research Theme(s) quoted and then relevant Topic(s).
Cell Envelope of Mycobacterium tuberculosis: biogenesis and role in virulence
Secondary Supervisor(s): Professor Gurdyal Besra
University of Registration: University of Birmingham
BBSRC Research Themes: Understanding the Rules of Life (Immunology, Microbiology)
Project Outline
Background: The genome of Mycobacterium tuberculosis, the causative agent of tuberculosis in humans, encodes a number of genes dedicated to lipid biosynthesis and transport. These unique lipids confer a distinct quality to the mycobacterial cell wall, and play an important role in biofilm formation, adhesion, immunomodulation and virulence. Research in the lab in the past has focused on deciphering the genetic pathways that define lipid biosynthesis and transport in pathogenic mycobacteria. We now make use of systems-based approaches to outline regulatory pathways that drive growth and cell wall biogenesis. With the use of defined mutant strains, we plan to test the molecular mechanisms underlying these processes, and their impact on virulence using experimental models of infection. Specifically, the project will focus on two key virulence lipids, mycolic acid and phthiocerol dimycocerosate (PDIM). Biogenesis and transport pathways for the former are also essential and a promising target for future drug development.
Objectives
1) Decipher molecular mechanisms underlying the regulation of mycolic acid biosynthesis by MadR, a transcription repressor.
2) Study how PDIMs are transported across the mycomembrane by a combination of fluorescence microscopy and structural studies on candidate proteins.
3) Develop an ex vivo lung infection model for studying the immune response to mycobacterial virulence lipids.
The experimental approaches taken to achieve the project objectives include:
1. Generation of gene knockouts and recombinant strains of pathogenic mycobacteria.
2. Protein expression and purification for Cryo EM based elucidation of structure.
3. Biophysical characterisation of proteins.
4. Biochemical analysis methodology including TLC, Mass Spectroscopy and NMR.
5. RNA Seq studies on mycobacterial transcription factor mutants.
6. Fluorescence microscopy of mycobacteria.
7. Tissue culture methodology applied to ex vivo tissue samples.
References
1. Cooper C, Peterson EJR, Bailo R, Pan M, Singh A, Moynihan P, Nakaya M, Fujiwara N, Baliga N, Bhatt A (2022) MadR mediates acyl CoA-dependent regulation of mycolic acid desaturation in mycobacteria. Proc Natl Acad Sci 119:e2111059119.
2. Moolla N, Bailo R, Marshall R, Bavro VN, Bhatt A. (2021) Structure-function analysis of MmpL7-mediated lipid transport in mycobacteria. Cell Surf 7:100062.
