Mechanisms of Host Defense Against Membrane Damage by Pore-Forming Toxins

Membrane damage is a common occurrence during microbial infection, but host signaling pathways responsive to this damage are poorly understood. Moreover, the mechanisms that promote membrane integrity in response to bacterially-induced damage remain incompletely defined. Thus, the goal of this research is to elucidate host pathways that guard against cellular damage, which can contribute to disease pathology during infection and inflammation. The objective of the proposed work is to define specific regulators of cellular defense again membrane damage by microbial pore- forming toxins (PFT). We have identified the cysteine aspartate protease, caspase-7, as a pivotal regulator of the macrophage response to pore formation by bacterial toxins. Caspase-7 activation is triggered during Listeria monocytogenes infection by the cholesterol dependent cytolysin, Listeriolysin O (LLO). Caspase-7 activation is also induced by sublethal concentrations of recombinant purified PFT, but not by sublethal concentrations of detergents. These data suggest that caspase-7 activation is selectively responsive to biological membrane-damaging agents. Finally, our preliminary data reveal a requirement for caspase-7 in maintaining host membrane integrity during Listeria infection. <p/>Previous studies had established caspase-7 as an effector of programmed cell death, but the protective effect of caspase-7 during L. monocytogenes infection occurs independently of apoptosis and does not require key adaptors of the TLR, NLR or inflammasome pathways. Thus, we find that caspase-7 plays a novel cytoprotective role during bacterial infection of macrophages. <p/>We therefore hypothesize that pore formation by bacterial toxins induces membrane repair mechanisms in a caspase-7 dependent manner. We will test this hypothesis in the following specific aims: (1) Define the nature of the bacterial membrane damage signal that triggers caspase-7 activation; (2) Identify regulators of caspase-7 induction in response to pore formation; (3) Determine the mechanism of caspase-7 dependent membrane repair and its contribution to host responses in vivo. Investigating mechanisms underlying the caspase-7 dependent cytoprotective response will provide fundamental insights into cellular function relevant to many aspects of disease. The expected outcome of this work will be to define a novel cellular function for caspase-7 during infection by bacterial pathogens. In addition to their role in bacterial pathogenesis, mammalian pore-forming proteins also participate in the immune response. Thus, mechanisms by which mammalian cells protect themselves from membrane damage may be relevant to protection from destruction by immune mechanisms as well as microbial toxins. The impact of the proposed research will be to elucidate a previously unappreciated damage control mechanism that could provide insight into future therapeutic approaches to minimizing cell and tissue damage in pathological disease states.