Identification and characterization of new antimicrobial host-factors:

Activation of the mammalian innate immune system by IFNγ is essential for host resistance to most pathogens. Cell-autonomous immunity to bacterial pathogens (e.g., Chlamydia trachomatis) and protozoan pathogens (e.g., Toxoplasma gondii) is controlled by two families of IFN-inducible GTPases: Immunity Related GTPases (IRGs) and Guanylate binding proteins (GBPs). Members of these two GTPase families associate with PVs and solicit antimicrobial resistance pathways specifically to the intracellular site of infection. The effector functions of IRG and GBP proteins are poorly defined but are believed to be mediated through the recruitment of additional host defense proteins at the site of infection. With a unique proteomics approach a bunch of effector proteins was identified that includes E3-ubiquitin ligases which specifically catalyze the ubiquitination of target proteins. One of our lab research is to characterize the function of such effector proteins, including E3-ubiquitin ligases, as host resistant factors in cell-autonomous immunity against aforesaid intravacuolar pathogens. Our current work is geared towards the development of a detailed molecular understanding of host-mediated ubiquitination of PVs which help to detect and destroy the PVs through Xenophagy (a selective type of autophagy to eliminate pathogens) in both mouse and human cells. To study this pivotal aspect of host-pathogen interaction, we are taking molecular, cellular and biochemical approaches alongside with various genetic tools such as CRISPR/Cas9-mediated genome editing.

Understanding of the counteracting immune-evasion strategies by intracellular pathogens against host ubiquitin-immune recognition of pathogen containing vacuoles (PVs):

Since ubiquitination and deubiquitination regulate several essential cellular processes, such as protein degradation, cell-cycle progression, DNA repair and signaling in innate and adaptive immunity, it is not surprising that many microbes have developed the means to interfere various ubiquitination pathways to promote their survival and replication. Recent studies demonstrated that ubiquitination of PVs (containing C. trachomatis or T. gondii) is advantageous to the host and that hyper-virulent strains of these pathogens have evolved strategies to interfere with IFNγ-inducible PV ubiquitination pathways. Mechanisms for pathogen evasion of IFNγ-inducible PV ubiquitination pathways are still poorly understood. Defining these pathways on a molecular level and identifying the microbial evasion mechanisms may reveal novel microbial targets. We predict that highly virulent pathogens such as Chlamydia trachomatis, T. gondii (Type-I) and drug resistant Leishmania spp are equipped with unique virulence factors to subvert PV ubiquitination pathways and establish chronic/persistent infections. One of our research goals is to identify novel ‘anti-ubiquitination’ virulence factors expressed by Leishmania spp and specific strains of T. gondii that are naturally resistant to host-mediated PV ubiquitination. Subsequently we’ll also expand our research to identify and characterize anti-ubiquitinating virulence factor(s) expressed by multi-drug resistant Mycobacterium spp. We will use biochemical, proteomics and genetic approaches to identify microbial virulence factors as well as host factors that are exploited by the pathogens to block or reverse PV ubiquitination.

Ultimately, we hope an increased understanding of pathogen ubiquitin immune-evasion strategies will help us to discover novel microbial targets and this will contribute to the development of more effective, alternative therapeutic strategies to treat microbial infections particularly in drug-resistant cases.