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The goal of this research is to assess the risk of humans to mycotoxins via foodborne and airborne exposure and develop appropriate mitigation strategies to protect human and animal health. The two primary objectives of this project are to: <OL> <LI> Determine the mechanisms by which simple trichothecenes aberrantly modulate immune function and impair weight gain <LI>Understand the molecular basis by which macrocylclic trichothecenes kill olfactory sensory neurons.
NON-TECHNICAL SUMMARY: We propose to learn how a class of potent biological toxins interferes with the function and survival of cells that are essential to the immune and nervous systems. This research will enhance our capacity to assess and manage risks associated with exposure to these toxins as well as yield mechanism-based strategies for preventing and/or treating persons exposed to these agents via inadvertant food/air contamination or deliberate chemical terrorism. Collectively, these outcomes will positively impact public health by providing a scientific basis for generating sound recommendations relative to these important toxins and appropriate remedial actions should exposures occur.
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APPROACH: In Objective 1, we will test the hypothesis that the ribosome plays a central role in the initiation and integration of protein kinase-mediated stress responses to trichothecenes and other ribotoxic agents. This hypothesis is based on observations in the macrophage that: 1) deoxynivalenol (DON), a common foodborne trichothecene, mediates cleavage of 18S and 28S ribosomal (r)RNA, 2) double-stranded RNA-activated protein kinase (PKR), a ribosome-associated serine-threonine kinase, is essential for DON-induced protein kinase activation and 3) DON induces mobilization of several protein kinases to the ribosome whereupon they are phosphorylated. We will test our hypothesis by: (1) characterizing DON-induced rRNA cleavage relative to targets, kinetics and mechanisms in the macrophage; (2) using both macrophage and cell-free models to characterize the role of PKR as an early sensor of DON-induced rRNA damage, (3) tracking DON-induced changes in ribosome-associated proteins in the macrophage relative to composition, kinetics and kinase activities; and (4) confirming putative mechanisms uncovered in in vitro studies in the mouse. From these studies, we expect to understand how the ribosome mediates the induction and integration of multiple intracellular signaling cascades that drive altered gene expression and apoptosis in mononuclear phagocytes in response to ribotoxic agents. In Objective 2, we will test the hypothesis that the macrocyclic trichothecene satratoxin G(SG) induces both rapid expression of death receptor genes (TNF-á, FasL and TRAIL) as well as the rapid degradation of a critical Bcl-2 family anti-apoptotic protein, Mcl-1 in OP6 cells, a clonal OSN model. This hypothesis is based on our observations that SG induces both rapid expression of death receptor genes (TNF-á, FasL and TRAIL) as well as the rapid degradation of a critical Bcl-2 family anti-apoptotic protein, Mcl-1 in OP6 cells, a clonal OSN model. We will test our hypothesis by: (1)characterizing roles of death receptor ligands in the induction of translation-dependent apoptosis by SG in OP6 cells; (2) determining contribution of anti-apoptotic protein instability in the induction of translation-independent apoptosis by SG in OP6 cells; (3)characterizing translation-dependent and translation-independent mechanisms for SG-induced OSN apoptosis in primary mouse olfactory epithelium cultures ; and (4) relating translation-dependent and translation-independent mechanisms to SG-induced OSN apoptosis and olfactory epithelium atrophy in the mouse.