Dataset Information

FOXO1 represses sprouty 2 and sprouty 4 expression to promote arterial specification and vascular remodeling in the mouse yolk sac.

ABSTRACT: Establishing a functional circulatory system is required for post-implantation development during murine embryogenesis. Previous studies in loss-of-function mouse models showed that FOXO1, a Forkhead family transcription factor, is required for yolk sac (YS) vascular remodeling and survival beyond embryonic day (E) 11. Here, we demonstrate that at E8.25, loss of Foxo1 in Tie2-cre expressing cells resulted in increased sprouty 2 (Spry2) and Spry4 expression, reduced arterial gene expression and reduced Kdr (also known as Vegfr2 and Flk1) transcripts without affecting overall endothelial cell identity, survival or proliferation. Using a Dll4-BAC-nlacZ reporter line, we found that one of the earliest expressed arterial genes, delta like 4, is significantly reduced in Foxo1 mutant YS without being substantially affected in the embryo proper. We show that FOXO1 binds directly to previously identified Spry2 gene regulatory elements (GREs) and newly identified, evolutionarily conserved Spry4 GREs to repress their expression. Furthermore, overexpression of Spry4 in transient transgenic embryos largely recapitulates the reduced expression of arterial genes seen in conditional Foxo1 mutants. Together, these data reveal a novel role for FOXO1 as a key transcriptional repressor regulating both pre-flow arterial specification and subsequent vessel remodeling within the murine YS.

SUBMITTER: Li-Villarreal N

PROVIDER: S-EPMC8995087 | biostudies-literature | 2022 Apr

REPOSITORIES: biostudies-literature

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FOXO1 represses sprouty 2 and sprouty 4 expression to promote arterial specification and vascular remodeling in the mouse yolk sac.

Li-Villarreal Nanbing N Wong Rebecca Lee Yean RLY Garcia Monica D MD Udan Ryan S RS Poché Ross A RA Rasmussen Tara L TL Rhyner Alexander M AM Wythe Joshua D JD Dickinson Mary E ME

Development (Cambridge, England) 20220401 7

Establishing a functional circulatory system is required for post-implantation development during murine embryogenesis. Previous studies in loss-of-function mouse models showed that FOXO1, a Forkhead family transcription factor, is required for yolk sac (YS) vascular remodeling and survival beyond embryonic day (E) 11. Here, we demonstrate that at E8.25, loss of Foxo1 in Tie2-cre expressing cells resulted in increased sprouty 2 (Spry2) and Spry4 expression, reduced arterial gene expression and r ...[more]

PMID: 35297995

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Project description:BackgroundThe signaling cascades that direct the morphological differentiation of the vascular system during early embryogenesis are not well defined. Several signaling pathways, including Notch and VEGF signaling, are critical for the formation of the vasculature in the mouse. To further understand the role of Notch signaling during endothelial differentiation and the genes regulated by this pathway, both loss-of-function and gain-of-function approaches were analyzed in vivo.ResultsConditional transgenic models were used to expand and ablate Notch signaling in the early embryonic endothelium. Embryos with activated Notch1 signaling in the vasculature displayed a variety of defects, and died soon after E10.5. Most notably, the extraembryonic vasculature of the yolk sac displayed remodeling differentiation defects, with greatly enlarged lumens. These phenotypes were distinct from endothelial loss-of-function of RBPJ, a transcriptional regulator of Notch activity. Gene expression analysis of RNA isolated from the yolk sac endothelia of transgenic embryos indicated aberrant expression in a variety of genes in these models. In particular, a variety of secreted factors, including VEGF and TGF-β family members, displayed coordinate expression defects in the loss-of-function and gain-of-function models.ConclusionsMorphological analyses of the in vivo models confirm and expand the understanding of Notch signaling in directing endothelial development, specifically in the regulation of vessel diameter in the intra- and extraembryonic vasculature. Expression analysis of these in vivo models suggests that the vascular differentiation defects may be due to the regulation of key genes through the Notch-RBPJ signaling axis. A number of these genes regulated by Notch signaling encode secreted factors, suggesting that Notch signaling may mediate remodeling and vessel diameter in the extraembryonic yolk sac via autocrine and paracrine cell communication. We propose a role for Notch signaling in elaborating the microenvironment of the nascent arteriole, suggesting novel regulatory connections between Notch signaling and other signaling pathways during endothelial differentiation.

Project description:BackgroundSerum response factor (SRF), a member of the MADS box family of nuclear transcription factors, plays an important role in cardiovascular development and function. Numerous studies demonstrate a central role for SRF in regulating smooth and cardiac muscle cell gene expression. Consistent with this, loss of SRF function blocks differentiation of coronary vascular smooth muscle cells from proepicardial precursors, indicating SRF is necessary for coronary vasculogenesis. The role of SRF in endothelial cell contribution during early vascular development, however, has not been addressed. To investigate this, we generated transgenic mice lacking expression of SRF in endothelial cells. Mice expressing Cre recombinase (Tie2Cre+) under Tie2 promoter control were bred to mice homozygous for Srf alleles containing loxP recombination sites within the Srf gene (Srff/f). Tie2 is a tyrosine kinase receptor expressed predominantly on endothelial cells that mediates signalling during different stages of blood vessel remodelling. Resulting embryos were harvested at specific ages for observation of physical condition and analysis of genotype.ResultsTie2Cre+/-Srff/f embryos appeared to develop normally compared to wild-type littermates until embryonic day 10.5 (E10.5). Beginning at E11.5, Tie2Cre+/-Srff/f embryos exhibited cerebrovascular hemorrhaging and severely disrupted vascular networks within the yolk sac. Hemorrhaging in mutant embryos became more generalized with age, and by E14.5, most Tie2Cre+/-Srff/f embryos observed were nonviable and grossly necrotic. Hearts of mutant embryos were smaller relative to overall body weight compared to wild-type littermates. Immunohistochemical analysis revealed the presence of vascular endothelial cells; however, vessels failed to undergo appropriate remodelling. Initial analysis by electron microscopy suggested a lack of appropriate cell-cell contacts between endothelial cells. Consistent with this, disrupted E-cadherin staining patterns were observed in mutant embryos.ConclusionThese results provide the first in vivo evidence in support of a role for SRF in endothelial cell function and strongly suggest SRF is required for appropriate vascular remodelling.

Dataset Information

FOXO1 represses sprouty 2 and sprouty 4 expression to promote arterial specification and vascular remodeling in the mouse yolk sac.

Publications

FOXO1 represses sprouty 2 and sprouty 4 expression to promote arterial specification and vascular remodeling in the mouse yolk sac.

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