MAM Proteins in Lung Vascular Injury

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Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) is a life-threatening condition which affects ~200,000 Americans each year with 10-15% of intensive care admissions and a mortality rate of ~ 25-40%. Sepsis is a prominent extrapulmonary cause of ALI in humans. Nearly 50% of patients with severe sepsis develop ALI/ARDS. All current therapies for ALI/ARDS still rely on supportive care; thus, there is an urgent need to develop new treatment strategies for ALI/ARDS that are safe and effective. Because endothelial cell (EC) barrier dysfunction is an early and critical component of ALI in sepsis, a better understanding of the mechanisms of EC permeability is key to developing effective therapy for ALI. The goal of this proposal is to understand how aberrant communication between endoplasmic reticulum (ER) and mitochondrion in sepsis, mediated by ER chaperone BiP/GRP78 and mitochondrial (MITO) chaperone Mortalin/GRP75, promotes EC barrier dysfunction to cause ALI. Our hypothesis is that a close interaction and functional cooperation between BiP/GRP78 and Mortalin/GRP75 at the MAMs (mitochondrial associated endoplasmic reticulum membranes) serves to increase cytosolic Ca2+ and ER-MITO contact sites to cause mitochondrial Ca2+ overload and inflammasome activation, leading to lung vascular injury in sepsis. We further hypothesize that targeting BiP/GRP78 and Mortalin/GRP75 in combination may prove a superior therapeutic approach against sepsis-induced ALI and mortality. The proposal is based on our novel findings that BiP/GRP78 and Mortalin/GRP75 are key regulators of Ca2+ signaling and ER-MITO contact sites in EC and silencing BiP/GRP78 or Mortalin/GRP75 each prevents EC barrier disruption caused by plasma from septic patients and other clinically relevant edemagenic agonists such as thrombin and LPS. BiP/GRP78 is induced in septic lung and overexpressing BiP/GRP78 in the resting lung endothelium is sufficient to cause lung injury in mice. Moreover, inhibiting BiP/GRP78 or Mortalin/GRP75 each mitigates ALI in mice. Importantly, combined inhibition of BiP/GRP78 or Mortalin/GRP75 is far more effective (requires ~5-fold less dose of BiP/GRP78 or Mortalin/GRP75 inhibitors) in improving survival in mice with sepsis. The proposal will address the following aims. Aim 1 will determine the role of BiP/GRP78 and Mortalin/GRP75 in regulating cytosolic Ca2+ to cause EC permeability. Aim 2 will determine the role of BiP/GRP78 and Mortalin/GRP75 in increasing ER-MITO contact sites and mitochondrial Ca2+ uptake to cause mitochondrial dysfunction and inflammasome activation leading to EC permeability. Aim 3 will determine (i) the contribution of endothelial BiP/GRP78 and Mortalin/GRP75 in causing lung injury, and (ii) assess the therapeutic potential of inhibiting BiP/GRP78 and Mortalin/GRP75 together against sepsis-induced ALI and mortality. These studies will provide valuable insight into the ER-MITO communication and its relevance in the pathogenesis of ALI and may lead to development of a safe and efficacious therapeutic strategy that relies on combined inhibition of BiP/GRP78 and Mortalin/GRP75 to control ALI and improve survival in sepsis.