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Title:Characterization of the Helicobacter pylori vacuolating cytotoxin mediated induction of mitochondrial cell death mechanism
Author(s):Jain, Prashant
Director of Research:Blanke, Steven R.
Doctoral Committee Chair(s):Blanke, Steven R.
Doctoral Committee Member(s):Salyers, Abigail A.; Slauch, James M.; Whitaker, Rachel J.
Department / Program:Microbiology
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Helicobacter pylori
Helicobacter pylori (VacA)
Cell death
Mitochondrial fission
Abstract:The Helicobacter pylori vacuolating cytotoxin (VacA) is a paradigm for microbial virulence factors that target and disable host cell mitochondria. VacA intoxication triggers mitochondrial dysfunction and metabolic stress, but the mechanisms underlying these perturbations in mitochondrial function are poorly understood. In this dissertation, we report that infection of gastric epithelial cells with H. pylori results in the transition of cellular mitochondria from a predominantly filamentous network state to small punctiform organelles, suggesting that H. pylori uncouples the dynamic balance between mitochondrial fusion and division that normally exists within healthy cells. H. pylori infection induces excessive mitochondrial recruitment of the endogenous host protein Drp1, which plays a critical role in the regulation of mitochondrial division within uninfected healthy cells. The dynamic balance between mitochondrial division and fusion is important for maintaining proper functionality of the network. However, increased mitochondrial fragmentation could result in metabolic stress and cell death. Among H. pylori factors, the vacuolating cytotoxin (VacA) was demonstrated to be both essential and sufficient to disrupt mitochondrial dynamics. Importantly, specific inhibition of Drp1 activity blocks VacA-induced fragmentation of mitochondrial network, as well as activation of mitochondria dependent cell death program (apoptosis), a hallmark of H. pylori infection. Importantly, Drp1 mediated mitochondrial fission preceded and was required for Bax activation, which is critical for VacA dependent cell death mechanism. However, the complete mechanism underlying VacA mediated Bax activation, particularly the nature of cellular changes initiated by VacA that linked the de-regulation of mitochondrial network dynamics to engagement of host apoptotic machinery, was not entirely clear. Additional studies reported in Chapter 3 of this dissertation demonstrate that VacA induces the activation and mitochondrial recruitment of the endogenous stress sensor protein Bid (BH3 interacting death domain agonist). Within VacA intoxicated cells, Bid was important for the activation of Bax and mitochondrial cell death mechanism. Importantly, Drp1 GTPase activity was required for Bid activation within VacA intoxicated cells. Our results therefore indicate that cellular stress as a result of excessive fission is possibly translated to Bax mediated mitochondrial outer membrane permeabilization through the stress sensor Bid. Furthermore, Drp1 dependent mitochondrial fission also resulted in the increase in cytosolic calcium levels, which was required for the activation of the calcium dependent cysteine protease called calpain. The proteolytic activity of calpain was required for processing of Bid to yield the mitochondrial targeting active fragment called t-Bid (truncated Bid). We Hypothesize that H. pylori mediated deregulation of mitochondrial dynamics promotes bacterial colonization by generating a novel class of dysfunctional host cells that are energetically crippled, and unable to respond appropriately to infection with H. pylori at the epithelial barrier.
Issue Date:2012-06-27
Rights Information:Copyright 2012 Prashant Jain
Date Available in IDEALS:2012-06-27
Date Deposited:2012-05

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