The vascular endothelial growth factor (VEGF) isoform VEGF165 stimulates vascular growth and hyperpermeability. Whereas blood vessel growth is essential to sustain organ health, chronic ...hyperpermeability causes damaging tissue edema. By combining in vivo and tissue culture models, we show here that VEGF165-induced vascular leakage requires both VEGFR2 and NRP1, including the VEGF164-binding site of NRP1 and the NRP1 cytoplasmic domain (NCD), but not the known NCD interactor GIPC1. In the VEGF165-bound receptor complex, the NCD promotes ABL kinase activation, which in turn is required to activate VEGFR2-recruited SRC family kinases (SFKs). These results elucidate the receptor complex and signaling hierarchy of downstream kinases that transduce the permeability response to VEGF165. In a mouse model with choroidal neovascularisation akin to age-related macular degeneration, NCD loss attenuated vessel leakage without affecting neovascularisation. These findings raise the possibility that targeting NRP1 or its NCD interactors may be a useful therapeutic strategy in neovascular disease to reduce VEGF165-induced edema without compromising vessel growth.
Visual information is relayed from the eye to the brain via retinal ganglion cell (RGC) axons. Mice lacking NRP1 or NRP1-binding VEGF-A isoforms have defective RGC axon organisation alongside brain ...vascular defects. It is not known whether axonal defects are caused exclusively by defective VEGF-A signalling in RGCs or are exacerbated by abnormal vascular morphology. Targeted NRP1 ablation in RGCs with a
knock-in allele reduced axonal midline crossing at the optic chiasm and optic tract fasciculation. In contrast,
-mediated endothelial NRP1 ablation induced axon exclusion zones in the optic tracts without impairing axon crossing. Similar defects were observed in
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mice, which have vascular defects as a result of their expression of single VEGF-A isoforms. Ectopic midline vascularisation in endothelial
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mutants caused additional axonal exclusion zones within the chiasm. As
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assays demonstrated that vessels do not repel axons, abnormally large or ectopically positioned vessels are likely to present physical obstacles to axon growth. We conclude that proper axonal wiring during brain development depends on the precise molecular control of neurovascular co-patterning.
Neuropilin 1 (NRP1) is expressed by neurons, blood vessels, immune cells and many other cell types in the mammalian body and binds a range of structurally and functionally diverse extracellular ...ligands to modulate organ development and function. In recent years, several types of mouse knockout models have been developed that have provided useful tools for experimental investigation of NRP1 function, and a multitude of therapeutics targeting NRP1 have been designed, mostly with the view to explore them for cancer treatment. This review provides a general overview of current knowledge of the signalling pathways that are modulated by NRP1, with particular focus on neuronal and vascular roles in the brain and retina. This review will also discuss the potential of NRP1 inhibitors for the treatment for neovascular eye diseases.
Neuropilin (NRP) 1 is a receptor for the vascular endothelial growth factor (VEGF)-A and is essential for normal angiogenesis. Previous in vitro experiments identified NRP1 interactions with VEGF-A's ...main signaling receptor VEGFR2 within endothelial cells, but also between nonendothelial NRP1 and endothelial VEGFR2. Consistent with an endothelial role for NRP1 in angiogenesis, we found that VEGFR2 and NRP1 were coexpressed in endothelial tip and stalk cells in the developing brain. In addition, NRP1 was expressed on two cell types that interact with growing brain vessels—the neural progenitors that secrete VEGF-A to stimulate tip cell activity and the pro-angiogenic macrophages that promote tip cell anastomosis. Selective targeting of Nrp1 in each of these cell types demonstrated that neural progenitor- and macrophage-derived NRP1 were dispensable, whereas endothelial NRP1 was essential for normal brain vessel growth. NRP1 therefore promotes brain angiogenesis cell autonomously in endothelium, independently of heterotypic interactions with nonendothelial cells. Genetic mosaic analyses demonstrated a key role for NRP1 in endothelial tip rather than stalk cells during vessel sprouting. Thus, NRP1-expressing endothelial cells attained the tip cell position when competing with NRP1-negative endothelial cells in chimeric vessel sprouts. Taken together, these findings demonstrate that NRP1 promotes endothelial tip cell function during angiogenesis.
•NRP1 promotes brain angiogenesis cell autonomously in endothelium, independently of heterotypic interactions with nonendothelial cells.•NRP1 plays a key role in endothelial tip rather than stalk cells during vessel sprouting in the brain.
Neuropilin 1 (NRP1) plays an important but ill-defined role in VEGF-A signaling and vascular morphogenesis. We show that mice with a knockin mutation that ablates the NRP1 cytoplasmic tail (Nrp1cyto) ...have normal angiogenesis but impaired developmental and adult arteriogenesis. The arteriogenic defect was traced to the absence of a PDZ-dependent interaction between NRP1 and VEGF receptor 2 (VEGFR2) complex and synectin, which delayed trafficking of endocytosed VEGFR2 from Rab5+ to EAA1+ endosomes. This led to increased PTPN1 (PTP1b)-mediated dephosphorylation of VEGFR2 at Y1175, the site involved in activating ERK signaling. The Nrp1cyto mutation also impaired endothelial tubulogenesis in vitro, which could be rescued by expressing full-length NRP1 or constitutively active ERK. These results demonstrate that the NRP1 cytoplasmic domain promotes VEGFR2 trafficking in a PDZ-dependent manner to regulate arteriogenic ERK signaling and establish a role for NRP1 in VEGF-A signaling during vascular morphogenesis.
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•The NRP1 cytoplasmic domain promotes VEGF receptor (VEGFR) 2 endocytic trafficking•In its absence, VEGR2 trafficking is delayed in sorting endosomes•PTP1b binds to Rab5+ sorting endosomes and dephosphorylates the Y1175 site of VEGFR2•Loss of the NRP1 cytoplasmic domain impairs developmental and adult arteriogenesis
Lanahan et al. find that the VEGF coreceptor NRP1 is required for proper VEGFR2 endocytic trafficking. In knockin mice lacking NRP1’s cytoplasmic domain, VEGFR2 passes through early endocytic compartments more slowly, increasing its exposure to the inhibitory phosphatase PTP1b and attenuating VEGF-stimulated ERK signaling and arteriogenesis.
Individuals with an inherited deficiency in gonadotropin-releasing hormone (GnRH) have impaired sexual reproduction. Previous genetic linkage studies and sequencing of plausible gene candidates have ...identified mutations associated with inherited GnRH deficiency, but the small number of affected families and limited success in validating candidates have impeded genetic diagnoses for most patients. Using a combination of exome sequencing and computational modeling, we have identified a shared point mutation in semaphorin 3E (SEMA3E) in 2 brothers with Kallmann syndrome (KS), which causes inherited GnRH deficiency. Recombinant wild-type SEMA3E protected maturing GnRH neurons from cell death by triggering a plexin D1-dependent (PLXND1-dependent) activation of PI3K-mediated survival signaling. In contrast, recombinant SEMA3E carrying the KS-associated mutation did not protect GnRH neurons from death. In murine models, lack of either SEMA3E or PLXND1 increased apoptosis of GnRH neurons in the developing brain, reducing innervation of the adult median eminence by GnRH-positive neurites. GnRH neuron deficiency in male mice was accompanied by impaired testes growth, a characteristic feature of KS. Together, these results identify SEMA3E as an essential gene for GnRH neuron development, uncover a neurotrophic function for SEMA3E in the developing brain, and elucidate SEMA3E/PLXND1/PI3K signaling as a mechanism that prevents GnRH neuron deficiency.
Sprouting blood vessels are led by filopodia-studded endothelial tip cells that respond to angiogenic signals. Mosaic lineage tracing previously revealed that NRP1 is essential for tip cell function, ...although its mechanistic role in tip cells remains poorly defined. Here, we show that NRP1 is dispensable for genetic tip cell identity. Instead, we find that NRP1 is essential to form the filopodial bursts that distinguish tip cells morphologically from neighboring stalk cells, because it enables the extracellular matrix (ECM)-induced activation of CDC42, a key regulator of filopodia formation. Accordingly, NRP1 knockdown and pharmacological CDC42 inhibition similarly impaired filopodia formation in vitro and in developing zebrafish in vivo. During mouse retinal angiogenesis, CDC42 inhibition impaired tip cell and vascular network formation, causing defects that resembled those due to loss of ECM-induced, but not VEGF-induced, NRP1 signaling. We conclude that NRP1 enables ECM-induced filopodia formation for tip cell function during sprouting angiogenesis.
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•NRP1 enables morphological, but not genetic, specialization of endothelial tip cells•NRP1 is essential for matrix-induced CDC42 activation in endothelial cells•NRP1 promotes filopodia extension and actin remodeling via CDC42•Endothelial NRP1 loss and CDC42 inhibition similarly impair angiogenesis
During angiogenesis, new blood vessel sprouts are led by filopodia-studded tip cells to sense environmental signals and enable directional migration. NRP1 is a tip cell protein that senses angiogenic cues. Fantin et al. now show that NRP1 promotes tip cell function by enabling CDC42 activation for filopodia formation and actin remodeling.
Vascular endothelial growth factor (VEGF-A) is a major regulator of blood vessel formation and function. It controls several processes in endothelial cells, such as proliferation, survival, and ...migration, but it is not known how these are coordinately regulated to result in more complex morphogenetic events, such as tubular sprouting, fusion, and network formation. We show here that VEGF-A controls angiogenic sprouting in the early postnatal retina by guiding filopodial extension from specialized endothelial cells situated at the tips of the vascular sprouts. The tip cells respond to VEGF-A only by guided migration; the proliferative response to VEGF-A occurs in the sprout stalks. These two cellular responses are both mediated by agonistic activity of VEGF-A on VEGF receptor 2. Whereas tip cell migration depends on a gradient of VEGF-A, proliferation is regulated by its concentration. Thus, vessel patterning during retinal angiogenesis depends on the balance between two different qualities of the extracellular VEGF-A distribution, which regulate distinct cellular responses in defined populations of endothelial cells.
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