Graduate Thesis Or Dissertation
 

The MAV_4644 dual-function channel protein with putative ADP-ribosyltransferase activity of Mycobacterium avium is required for virulence within host macrophages

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https://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/j67318326

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  • Mycobacterium avium subspecies hominissuis (MAH) is an opportunistic pathogen that is ubiquitous in the environment and often isolated from faucets and showerheads. MAH mostly infects humans with an underlying disease, such as chronic pulmonary disorder (COPD), cystic fibrosis (CF), or are immunocompromised, though infections in patients without concurrent disease are increasing in prevalence. MAH is resistant to many antibiotics due to the impermeability of its envelope and the protection of its intracellular niche inside host macrophages, making the infections difficult to treat. Host macrophages produce nitric oxide and reactive oxygen intermediates to kill phagocytosed bacteria. However, MAH replicates in nitric oxide producing cells and is less virulent when inducible nitric oxide synthesis is suppressed in a murine host. To investigate the relationship between MAH and nitric oxide, an in vitro model was developed in murine macrophages. We show that MAH grows in macrophages stimulated with IFN-g and producing nitric oxide. A transposon library screen of MAH for mutants incubated with nitric oxide donor indicated that inactivation of the gene MAV_4644 (MAV_4644:Tn) resulted in nitric oxide susceptibility. Characterization of MAV_4644:Tn revealed that it is required for virulence in host macrophages. In silico analysis revealed MAV_4644 is a dual-function channel protein with putative ADP-ribosyltransferase activity. Protein binding assays indicate that MAV_4644 protein interacts with the host lysosomal peptidase, cathepsin Z. Cathepsins are key regulators of inflammation and antigen presentation within immune cells. Pathogenic mycobacteria have been shown to suppress the action of other cathepsins to establish their intracellular niche. Knock-down of cathepsin Z in host macrophages rescued the attenuated phenotype of MAV_4644:Tn. The data suggests cathepsin Z is involved in early mycobacterial killing within host macrophages and virulence factor MAV_4644 protein abrogates this process.
  • Keywords: mycobacterium avium hominissuis, macrophage, nitric oxide, cathepsin Z, MAV_4644
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