Abstract 5317: Bone Morphogenetic Protein Recruits a Peroxisome Proliferator-Activated Receptor γ and β-catenin Transcription Factor Complex to Regulate Apelin Expression in Pulmonary Endothelial Cells
Our previous studies on the pathobiology of pulmonary arterial hypertension (PAH) demonstrated that bone morphogenetic protein receptor II (BMPRII) mediated signaling plays a key role in pulmonary arterial (PA) endothelial cell (EC) survival and proliferation via activation of beta-catenin (βC). We have also shown that a βC interacting nuclear receptor, peroxisome proliferator-activated receptor γ (PPARγ), is a mediator of BMPRII signaling in PA smooth muscle cells (SMCs). A mouse model with targeted disruption of PPARγ in EC shows PAH. The downstream gene targets of BMPRII signaling via PPARγ in EC are, however, unknown. We hypothesized that BMPRII induces a PPARγ-βC transcription factor complex that regulates genes necessary for PAEC homeostasis. We first performed co-immunoprecipitation (IP) and confocal microscopy to demonstrate a relationship between formation of the PPARγ-βC complex and EC survival. We identified target genes for PPARγ-βC complex in PAEC by comparing those on whole genome wide ChIP-Chip analyses with those altered on expression profiling microarrays after silencing either BMPRII or βC by siRNA. PPARγ-βC complex targets were then analyzed by western immunoblotting and qPCR in PAEC from idiopatic PAH (IPAH) vs. control patients, and in PPARγ deficient vs. wild type mice. By IP and confocal microscopy we demonstrated that PPARγ interacts, and co-localizes in the nucleus, with βC in PAEC. This interaction was increased 3 fold upon BMP2 stimulation. Furthermore, ChIP-Chip analysis revealed βC co-occupancy in over 70% of PPARγ bound promoters. Microarray analyses confirmed significant changes in the expression of 35 of those genes after silencing of BMPRII and βC in PAEC. We then focused on apelin as a ChIP-Chip identified target important in endothelial homeostasis. Loss-of either BMPRII or βC down-regulated apelin > 1.5 fold, and apelin mRNA and protein were decreased in PAEC of IPAH vs. control patients (60% and 35% respectively) and in PPARγ deficient vs. WT mice (P < 0.05 for all). We conclude that BMP signaling induces a PPARγ-βC interaction that promotes EC homeostasis by regulating critical targets such as apelin. We speculate that down-regulation of apelin may be important in the pathobiology of PAH.