Hemolysis is one of the critical pathogenic mechanisms of pulmonary hypertension (PH), predisposing individuals with hemolytic conditions to PH. Notably, 10-30% of sickle cell disease patients and up to 80% of thalassemia patients develop PH, starkly contrasting to a mere 0.001% prevalence in the general population. Moreover, our recent findings indicate significant correlations between elevated sub-clinical hemolysis in Group 1 PAH patients and PAH severity. However, the critical mechanism of heme actions leading to PH progression still needs to be understood. By elucidating the impact of heme on the pulmonary vasculature, we aim to shed light on the contribution of heme- mediated mechanisms to the pathogenesis of Group 1 PAH. Our previous studies demonstrated that the free heme directly triggers intracellular signaling in endothelial cells by activating the MKK3/p38 pathway. However, the precise mechanism by which free heme activates this signaling axis remains to be determined. Our search for heme-sensor factors identified a Heme-Activating Protein (HAP1) responsible for sensing the heme and initiating heme target genes in Yeast. The homolog of HAP1 in humans is the INAVA, known to be responsible for triggering an immune response in macrophages and disrupting the barrier in epithelial cells. However, there is a gap in knowledge on the molecular mechanism of INAVA action in endothelial cells and its involvement in hemolytic complications and PH pathogenesis. Based on our compelling preliminary data, we introduce INAVA as a novel intracellular heme-sensor protein activating MKK3/p38-mediated signaling by engaging 14-3-3 kinase. Notably, both INAVA and 14-3-3 are overexpressed in the lung tissues of PAH patients and have increased interaction, implying a disease- related signaling role. We aim to examine the hypothesis that free heme, by binding to INAVA, induces 14-3-3 activation in the cytosol. This increases endothelial proliferation, barrier dysfunction, and cytokines, leading to PH pathogenesis. Our research effort, focused on investigating endothelial cell heterogeneity using a single-cell approach, allowed us to discern the distinct pulmonary endothelial cell populations, each possessing unique gene signatures and functional arrays. We found five novel populations of general capillary ECs, named gCapA-E, in addition to well-known pulmonary arterial, venous, lymphatic endothelium, and aerocytes. Based on our preliminary data, we hypothesize that heme possesses a differential effect on each of these endothelial cell populations, compromising the maintenance of adequate endothelial barrier by gCapA, impairing angiogenic properties of gCapC, and altering proliferation of gCapB.