Signaling mechanisms by which mitochondria regulates fibrosis in the lung

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 DESCRIPTION (provided by applicant): Despite the fact that Idiopathic Pulmonary Fibrosis (IPF) is a fatal and progressive lung disease of unknown etiology whose prevalence dramatically increases with age, few new pathobiologic models have emerged. Repetitive injury of mitochondrial-enriched type II alveolar epithelial cells (AECII) appears to be a key triggering event that leads to fibrosis, linked to secretion of pro- fibrotic cytokines, and increased apoptosis. Although AECII containing approximately 50% of the lung mitochondrial mass, the role of mitochondria in IPF pathobiology is unknown. We recently discovered that AECII from human IPF lung have accumulation of dysmorphic and dysfunctional mitochondria associated with very low expression of the crucial protective protein involved in mitochondrial homeostasis, PTEN-induced putative kinase 1 (PINK1) (J. Clin. Invest 2015). Low expression of PINK1 is associated with ER stress and aging, leading to increased susceptibility to cell apoptosis and fibrosis. However, no information is available how ER stress regulates PINK1 expression and how loss of PINK1 activates pro-fibrotic responses. Our novel preliminary studies suggest i) that the ER stress induces ATF3, a transcriptional repressor that reduced PINK1 levels leads to opening of the mitochondria permeability transition pore (mPTP) that sequentially leads to mitochondrial depolarization, apoptosis and leakage of mtDNA, a damage associated molecular pattern (DAMP) that induces TGF-β expression, and iii) PINK1 inhibits mPTP opening by interaction with the adenine nucleotide transporter 2/3 (ANT2/3), involved in mitochondrial pore opening negatively regulates PINK1 gene expression, ii) Based on these observations, we hypothesize that ER stress transcriptionally inhibits expression of the indispensable mitochondrial molecule, PINK1, in AECII thereby leading to mitochondrial dysfunction, via opening of the mPTP, apoptosis, and release of a mitochondrial DAMP that induces pro-fibrotic responses. To test this hypothesis, we propose: (1) To determine if ER stress induces ATF3 that transcriptionally silences PINK1 gene expression that promotes mitochondrial dysfunction in the AECII (Aim 1). Here, we will use AECII with specific deletion and overexpression of ATF3 to analyze its contribution to PINK1 mRNA synthesis and its ability to impair mitochondrial homeostasis, and ATF3 knockout mice to study their susceptibility to lung fibrosis. In addition, we will interrogate the PINK1 promoter to determine core cis-acting elements regulated by ATF3. (2) To investigate if PINK1 deficiency alters the mitochondrial membrane permeability that drives apoptosis and promotion of pro-fibrotic responses. We hypothesize that PINK1 through a novel interaction represses ANT2/3, the latter promotes mitochondrial membrane permeability in AECII. We will evaluate how PINK1-ANT2/3 interaction is essential in limiting the opening of the mPTP, reducing apoptosis, restricting release of mitochondrial DAMPs, and inducing TGF-β expression. In addition, we will analyze whether a neo-substrate for PINK1 that amplifies its catalytic activity will have a preventive and/or therapeutic effect against PINK1 deficiency-associated mitochondria damage and lung fibrosis. Overall, these studies bring forth a unique molecular model linking PINK1-deficiency to mitochondrial dysfunction, ensuing extracellular release of DAMPs, and the profibrotic response that sets the stage for identifying novel therapeutic targets in ameliorating severity of fibrosis in IPF.