Pulmonary arterial hypertension (PAH) is a life-threatening disease with unmet medical needs. Currentlyavailable therapies fail to substantially reduce PAH progression and mortality which remains near 50% five yearsafter diagnosis. The cancer-like proliferation of the distal pulmonary arteries is the primary cause of increasedpulmonary vascular resistance leading to right heart failure. Recent studies highlighted a critical role of metabolicreprogramming in triggering pulmonary vascular remodeling. However the particular mechanistic link thatconnects the metabolic reprogramming with the uncontrolled proliferation of pulmonary vascular cells has notbeen established. Nevertheless the lack of this knowledge generates a critical barrier that prevents effectivetherapeutics that target vascular remodeling. During the previous grant cycle we showed that increasedoxidative stress in the PAH lungs in patients and animal models results in the nitration mediated Akt activation.The activation of Akt via nitration of tyrosine Y350 induces overexpression of Pyruvate Carboxylase (PC) leadingto anaplerotic stimulation of remodeling. We reported that inhibition of both Akt nitration or PC-mediatedanaplerosis resulted in marked attenuation of PAH in preclinical models. In cell culture experiments we observedthat Akt nitration changes the pulmonary artery endothelial cells (PAEC) morphology proliferation rate and geneexpressions. The microarray profiling showed upregulation of multiple markers of Endothelial to MesenchymalTransition (EndMT) in response to Akt nitration (PDGFRa TGFbR SMAD3 RUNX2). To identify the possiblemechanisms of EndMT we performed a mass spectrometry analysis of PC interactome. We found a directbinding of PC to the Cyclin-Dependent Kinase 5 (CDK5) attenuated by Akt nitration inhibition. Our data indicatethat PC could activate CDK5 in the cytosol. CDK5 in turn phosphorylates RUNX2 a well-established mediatorof mesenchymal transition. Indeed two recent publications showed activation of CDKs and RUNX2 signaling inPAH patients. However these reports did not provide mechanistic insights. In the current proposal wehypothesize that Akt nitration triggers PC expression and accumulation in the cytosol leading to activation of theCDK5/RUNX2 axis-mediated EndMT and vascular remodeling in PAH. We will test this hypothesis with thefollowing aims: 1) To elucidate the role of nitration (Y350) mediated Akt activation in EndMT events; 2) Todetermine whether cytosolic PC plays a key role in CDK5/RUNX2 axis activation; 3) To examine the effect oftargeted protein degraders (PROTACs) on EndMT in vivo.