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Role of Mitochondrial Ca 2+ ‐Activated Cl − Channel anoctamin‐1 and mitochondrial fusion protein OPA1 in pulmonary arterial hypertension
Author(s) -
Cypress Michael W.,
Vang Alexander,
Fernandez-Nicolas Ana,
Mancini Thomas,
Jhun Bong Sook,
Clements Richard T.,
Choudhary Gaurav,
O-Uchi Jin
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.07091
Subject(s) - mitochondrion , mitochondrial apoptosis induced channel , microbiology and biotechnology , mitochondrial fusion , inner mitochondrial membrane , biology , membrane potential , chemistry , biochemistry , mitochondrial dna , gene
Activation of a Ca 2+ ‐activated chloride channel, anoctamin‐1 (Ano1), is associated with enhanced cell proliferation in multiple cell types, such as endothelial cells (ECs). Recent proteomics studies indicate that Ano1 may interact with mitochondrial proteins in addition to plasma membrane (PM)‐localized proteins. However, the expression levels of Ano1 and its functions in other cellular compartments including mitochondria remain unclear. We previously showed that Ano1 is overexpressed in pulmonary artery ECs isolated from patients with idiopathic pulmonary arterial hypertension (PAH) and that Ano1 activation promotes apoptosis of pulmonary artery ECs from patients with idiopathic PAH, likely via depolarization of mitochondrial membrane potential and increased mitochondrial reactive oxygen species (ROS) generation. Therefore, we hypothesized that Ano1 exists not only at PM, but also in the mitochondria, which may contribute to a hyperproliferative phenotype of ECs under PAH. Here, we determined the mitochondrial expression of Ano1 and further investigated its role in the mitochondria. Subcellular localization of Ano1 was determined by immunostaining of Ano1 in Rat lung microvascular ECs (RLMVECs) and by live cell imaging of HEK293T cells expressing GFP‐tagged Ano1. The mitochondrial and PM localization of Ano1 was also confirmed by biochemical assays using protein fractionation from RLMVECs. Moreover, co‐immunoprecipitation assay showed that Ano1 interacts with a mitochondrial fusion protein OPA1 which is the critical regulator of mitochondrial bioenergetics and cristae formation and possesses a trans‐membrane domain at the inner mitochondrial membrane (IMM). These results indicate that 1) Ano1 expresses at the IMM in addition to the PM and 2) Ano1 and OPA1 interactions may be partly involved in the molecular mechanism underlying the hyperproliferation and apoptosis resistance of pulmonary artery ECs during PAH