Importance of endothelial Hey1 expression for thoracic great vessel development and its distal enhancer for Notch-dependent endothelial transcription

Abstract

Thoracic great vessels such as the aorta and subclavian arteries are formed through dynamic remodeling of embryonic pharyngeal arch arteries (PAAs). Previous work has shown that loss of a basic helix-loop-helix transcription factor Hey1 in mice causes abnormal 4th PAA development and lethal great vessel anomalies resembling congenital malformations in humans. However, how Hey1 mediates vascular formation remains unclear. In this study, we revealed that Hey1 in vascular endothelial cells, but not in smooth muscle cells, played essential roles for PAA development and great vessel morphogenesis in mouse embryos. Tek-Cre-mediated Hey1 deletion in endothelial cells affected endothelial tube formation and smooth muscle differentiation in embryonic 4th PAAs and resulted in interruption of the aortic arch and other great vessel malformations. Cell specificity and signal responsiveness of Hey1 expression were controlled through multiple cis-regulatory regions. We found two distal genomic regions that had enhancer activity in endothelial cells and in the pharyngeal epithelium and somites, respectively. The novel endothelial enhancer was conserved across species and was specific to large caliber arteries. Its transcriptional activity was regulated by Notch signaling in vitro and in vivo, but not by ALK1 signaling and other transcription factors implicated in endothelial cell specificity. The distal endothelial enhancer was not essential for basal Hey1 expression in mouse embryos but may likely serve for Notch-dependent transcriptional control in endothelial cells together with the proximal regulatory region. These findings help understand significance and regulation of endothelial Hey1 as a mediator of multiple signaling pathways in embryonic vascular formation.

General Transcription Factor IIF Polypeptide 2: A Novel Therapeutic Target for Depression Identified Using an Integrated Bioinformatic Analysis

“…Several studies have recognized the importance of understanding the behaviors and functions of TFs ( Füllgrabe and Moore, 2018 ; Martínez Corrales and Alic, 2020 ). For example, HEY1, a transcription factor, is an effector molecule of the NOTCH signaling pathway ( Watanabe et al, 2020 ; Xie et al, 2020 ). In salivary adenoid cystic carcinoma, NOTCH is positively associated with HEY1 activation, and HEY1 affects the expression of NOTCH and contributes to cellular proliferation, apoptosis inhibition, and spheroid formation in vitro ( Xie et al, 2020 ).…”

NKL Homeobox Genes NKX2-3 and NKX2-4 Deregulate Megakaryocytic-Erythroid Cell Differentiation in AML

“…This physiological connection may be aberrantly captured by the related NKX2-4 in OCI-M2. Additional endothelial factors are encoded by FLI1 , HEY1 , IRF6 , and SOX7 [ 25 , 27 , 50 , 51 ]. Chromosomal amplifications and chromosomal gains underlie their aberrant activation in OCI-M2.…”

ETS‐dependent enhancers for endothelial‐specific expression of serum/glucocorticoid‐regulated kinase 1 during mouse embryo development

“…Genomic DNA fragments were inserted into pGL4.10 luciferase plasmid (Promega) containing the thymidine kinase promoter. Transfection of HEK293T cells was performed as we previously reported (Watanabe et al., 2020). Briefly, 100 ng of luciferase reporter plasmid, 50 ng of β‐galactosidase reporter plasmid and 1.5625–400 ng of effector plasmid were used per a well of 24‐well plates.…”

“…The amounts of effector plasmids in each experiment were described in Figure Legends. Luciferase and β‐galactosidase activities were measured as we previously reported (Watanabe et al., 2020). All assays were independently performed three times, which gave reproducible results.…”