*Purpose: Loss of endothelial cell (EC) identity is central to endothelial to mesenchymal transition (EndMT) implicated in neointimal hyperplasia in transplant vasculopathy. However, the molecular mechanisms counteracting this cell fate transition are not clearly understood. The DNA binding protein BCL6B has been shown to regulate angiogenesis, but otherwise nothing is known about its function. We aimed to determine the role of the endothelial-restricted transcriptional regulator, BCL6B, in maintenance of endothelial identity and potential role in EndMT.

*Methods: Public bulk and single cell transcriptome datasets from GEO, the Endothelial Atlas, Tabula Muris, Tabula Sapiens; and ChIP-Seq tracks from ENCODE, were analyzed for cell-type specific expression, histone modifications and BCL6B gene targets. In vitro, human aortic EC were treated with siRNA to GAPD or BCL6B, or with the HDAC inhibitor TSA, for 2-4 days, and gene expression was measured by RNA-Seq.

*Results: We identified that BCL6B, a transcriptional regulator, was highly restricted to and universally expressed in EC. Its expression was inducible by master transcriptional regulators controlling EC identity, correlated with canonical endothelial genes, and the BCL6B gene was directly bound by ERG and ETS1, critical maintenance factors in endothelium. Only endothelium but not 8 other cell types exhibited high levels of transcription, and active chromatin marks H3K27Ac, H3K4Me1, and H3K4Me3 at the BCL6B gene. Its expression was particularly enriched in an “angiogenic” subpopulation of EC in the heart, which were also characterized by genes implicated in VEGF signaling, cell cycle and vascular repair. On the other hand, BCL6B expression was lowest in endocardial endothelium, which have greater plasticity than other vascular beds to undergo EndMT and give rise to mesenchymal compartments during development. As a DNA binding protein, overexpressed BCL6B ChIP-Seq in non-endothelial cells showed significant peaks at numerous genes involved in endothelial specification and regulation of developmental processes, including LDLR, MEIS2, ROBO2, PROX1, ETV1, and MECOM. Knocking down BCL6B in human cardiac EC resulted in perturbation of genes involved in hypoxia, angiogenesis and TGFβ responses. Further, BCL6B was elevated during tubulogenesis, developmental angiogenesis and in response to VEGF; and downregulated upon knockdown of ERG/FLI1, endothelial dedifferentiation or treatment with an HDAC inhibitor, which also trigger EndMT.

*Conclusions: Our findings highlight a novel transcriptional regulator BCL6B, which is universally expressed in endothelium, for its putative function in specifying endothelial quiescence. We hypothesize that BCL6B maintains endothelial identity, and that loss of BCL6B expression promotes maladaptive vascular remodeling. Insights into the mechanisms of loss of endothelial resilience will yield knowledge applicable to transplant vasculopathy and more effective management of this and other occlusive vascular diseases.

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