carbamoyl‐phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (cad) regulates Notch signaling and vascular development in zebrafish

Abstract

Background: The interplay between Notch and Vegf signaling regulates angiogenesis in the embryo. Notch signaling limits the responsiveness of endothelial cells to Vegf to control sprouting. Despite the importance of this regulatory relationship, much remains to be understood about extrinsic factors that modulate the pathway. Results: During a forward genetic screen for novel regulators of lymphangiogenesis, we isolated a mutant with reduced lymphatic vessel development. This mutant also exhibited hyperbranching arteries, reminiscent of Notch pathway mutants. Positional cloning identified a missense mutation in the carbamoyl‐phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (cad) gene. Cad is essential for UDP biosynthesis, which is necessary for protein glycosylation and de novo biosynthesis of pyrimidine‐based nucleotides. Using a transgenic reporter of Notch activity, we demonstrate that Notch signaling is significantly reduced in cadhu10125 mutants. In this context, genetic epistasis showed that increased endothelial cell responsiveness to Vegfc/Vegfr3 signaling drives excessive artery branching. Conclusions: These findings suggest important posttranslational modifications requiring Cad as an unappreciated mechanism that regulates Notch/Vegf signaling during angiogenesis. Developmental Dynamics 244:1–9, 2015. © 2014 Wiley Periodicals, Inc.

SoxF factors induce Notch1 expression via direct transcriptional regulation during early arterial development

“…Whole-mount in situ hybridisation in zebrafish larvae was performed as described (Coxam et al, 2015;Duong et al, 2014). Section and whole-mount in situ hybridisation in mouse was performed as described (Metzis et al, 2013;Fowles et al, 2003).…”

Dynamic regulation of basement membrane protein levels promotes egg chamber elongation in Drosophila

“…S4A–C). Alternatively, recent work has suggested that SPARC may promote solubility of Col IV in the extracellular space (Pastor-Pareja and Xu, 2011; Shahab et al, 2015). In Drosophila larvae, Col IV is produced by the fat body and then distributed, via the hemolymph, to organs throughout the body.…”

“…The fat body itself is surrounded by a BM; thus, the Col IV produced by this organ must remain soluble in order to pass through this BM and diffuse to distant sites. Loss of SPARC in this system causes Col IV to accumulate around fat body cells in a cell-autonomous manner (Pastor-Pareja and Xu, 2011; Shahab et al, 2015). In contrast, Col IV produced by the follicle cells is meant to integrate into the adjacent BM immediately upon secretion.…”

“…Despite its importance in development and disease, the mechanism by which SPARC affects BM structure is uncertain. It has been proposed to regulate Collagen deposition, degradation, and adhesion to cells, but a coherent view of SPARC function is lacking (Chlenski et al, 2011; Harris et al, 2011; Martinek et al, 2008; Pastor-Pareja and Xu, 2011; Sage et al, 1989; Shahab et al, 2015). SPARC is expressed in early stage follicle cells and accumulates with Col IV in the BM (Fig.…”

mafba is a downstream transcriptional effector of Vegfc signaling essential for embryonic lymphangiogenesis in zebrafish

“…As ISH is insensitive at later developmental stages, we examined mafba expression in FACS (fluorescent-activated cell sorting)-sorted populations of arterial ECs (AECs), venous ECs (VECs), and LECs that we isolated from 60 hpf, 3 dpf, and 5 dpf embryos (Fig. 3B,C;Coxam et al 2014Coxam et al , 2015. Indicative of lineage-restricted expression in the vasculature, we found that mafba was highly enriched in VECs compared with AECs at early time points (comparable with known VEC markers) and was enriched in LECs compared with VECs by 5 dpf (comparable with known venous and lymphatic markers lyve1b and prox1a).…”