Understanding the Pathogenic Process of Tuberculosis

by way of Gene Inhibition

Supervisor. Dr. Nick West^*. Co-supervisor. Prof. Warwick Britton

*Mycobacterial Research Lab, Centenary Institute.

Ph. 9565 6163, email n.west@centenary.usyd.edu.au

Tuberculosis remains a global health burden of staggering proportions. Around 1 in 3 people are infected with Mycobacteria tuberculosis, the organism responsible for the disease, which kills 2 million people annually. The emergence of strains now resistant to almost all of our front line drugs has increased the urgency to develop the critical antibiotics required to eradicate the disease.

New drug targets need to be identified and this project will attempt to do this by inhibiting gene expression of major structural proteins and/or proteins of key signalling pathways. Specifically, antisense RNA molecules will be produced for these specialised targets and cloned into tightly regulated inducible systems. These constructs, once introduced into Mycobacterium bovis BCG, can be activated following infection of human macrophages. The impact of the resultant BCG gene knock-down, in an intracellular environment, will be analysed in terms of bacterial viability and macrophage response.

Within our laboratory, a global analysis of macrophage gene expression (microarray) following BCG infection identified many differentially regulated genes. This project will also investigate the effect of RNA interference of one of these specific macrophage (host) genes and its subsequent consequences following BCG infection.

This project will train students to become proficient in the following techniques:

Molecular biology - antisense RNA design, siRNA design, Polymerase Chain Reaction, gene cloning

Cell biology - mammalian cell culture, isolation of human macrophages, DNA transfection, macrophage infections and various assays of bacterial and cellular function.