Acute lung injury and acute respiratory distress syndrome (ALI/ARDS) are severe respiratory illnesses occurring after pneumonia or sepsis. Cell death is a hallmark of ALI/ARDS. Pseudomonas aeruginosa is a major bacterial isolate from pneumonia that rapidly develops multi-drug resistance in subjects who develop ALI/ARDS. The underlying mechanism (s) of how epigenetics affects the pathobiology of ARDS is unknown. Our exciting preliminary results demonstrate that a multifunctional chromatin modulator, Mortality factor 4 like 1 (Morf4l1) epigenetically induces cell death in response to P. aeruginosa infection. Morf4l1 protein levels are elevated in human pneumonia lung tissue and in lipopolysaccharide (LPS) treated or P. aeruginosa infected lung epithelial cells. Interestingly, Morf4l1 is a highly cytotoxic protein that is rapidly eliminated in cells by an orphan ubiquitin E3 ligase subunit calld Fbxl18. However, P. aeruginosa infection protects Morf4l1 from degradation by increasing Morf4l1 acetylation that antagonizes its ubiquitination and removal in cells. Overexpression of Morf4l1 leads to profound epithelial cell death and is lethal to mice. Preliminary data also suggest that P. aeruginosa infection enhances Morf4l1 nuclear entry that may allow it escape its cytosolic degradation to facilitate its epigenetic function to control inflammatory gene transcription. Depletion of Morf4l1 by shRNA in mice effectively represses P. aeruginosa induced cell death. Importantly, by in silico drug screening, we found an effective Morf4l1 antagonist, argatroban, an FDA approved anticoagulant currently in the market. Argatroban tightly binds to Morf4l1, inhibits its epigenetic function, reduces Morf4l1 mediated cell death, an promotes cell survival during P. aeruginosa infection. In addition, administration of argatroban improves survival, suppresses cell death, and ameliorates inflammation in experimental murine models of ALI. These data led to our hypothesis that Pseudomonas infection protects Morf4l1 from SCF-Fbxl18 mediated degradation through its subcellular relocation and modification, thereby epigenetically modulating inflammation and triggering cell death. As a corollary to this hypothesis, we will test if targeting Morf4l1 with an FDA approved drug may attenuate the severity of experimental ALI. We will test this hypothesis by executing two specific aims: (1): to determine if P. aeruginosa infection triggers signal transduction to activate lysine acetyltransferase Gcn5 to interact and catalyze Morf4l1 acetylation, and the acetylation increases Morf4l1 protein levels that protects it from E3 ubiquitin ligase SCFFbxl18 mediated ubiquitin-proteasomal proteolysis. (2): to identify if P. aeruginosa infection triggers Morf4l1 dephosphorylation that enhances Morf4l1 nuclear entry to leads to epigenetic changes and cell death. Last we will target Morf4l1 with a small molecule (argatroban) to attenuate acute lung injury. These studies will provide a significant conceptual advance regarding epigenetic regulation in the pathogenesis of ALI/ARDS. The project also provides new insight into the molecular control of a poorly described chromatin modulator (Morf4l1) in the regulation of cell death and its pathological role in ALI/ARDS. The study identifies an off-labeling effect of an FDA approved drug that paves a new way for pharmacotherapeutic targeting of Morf4l1 in ALI/ARDS pathogenesis.
P. aeruginosa infection is a highly virulent pathogen etiologically linked to the high morbidity and mortality from the acute lung injury and acute respiratory distress syndrome (ALI/ARDS), and in nosocomial pneumonia patients. These studies will identify novel molecular mechanisms on how P. aeruginosa infection epigenetically leads to host cell death. Based on these critical discoveries, we developed a small molecule directed at ameliorating acute inflammatory lung disease.
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