notch3is essential for oligodendrocyte development and vascular integrity in zebrafish

Abstract

SUMMARYMutations in the human NOTCH3 gene cause CADASIL syndrome (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy). CADASIL is an inherited small vessel disease characterized by diverse clinical manifestations including vasculopathy, neurodegeneration and dementia. Here we report two mutations in the zebrafish notch3 gene, one identified in a previous screen for mutations with reduced expression of myelin basic protein (mbp) and another caused by a retroviral insertion. Reduced mbp expression in notch3 mutant embryos is associated with fewer oligodendrocyte precursor cells (OPCs). Despite an early neurogenic phenotype, mbp expression recovered at later developmental stages and some notch3 homozygous mutants survived to adulthood. These mutants, as well as adult zebrafish carrying both mutant alleles together, displayed a striking stress-associated accumulation of blood in the head and fins. Histological analysis of mutant vessels revealed vasculopathy, including: an enlargement (dilation) of vessels in the telencephalon and fin, disorganization of the normal stereotyped arrangement of vessels in the fin, and an apparent loss of arterial morphological structure. Expression of hey1, a well-known transcriptional target of Notch signaling, was greatly reduced in notch3 mutant fins, suggesting that Notch3 acts via a canonical Notch signaling pathway to promote normal vessel structure. Ultrastructural analysis confirmed the presence of dilated vessels in notch3 mutant fins and revealed that the vessel walls of presumed arteries showed signs of deterioration. Gaps in the arterial wall and the presence of blood cells outside of vessels in mutants indicated that compromised vessel structure led to hemorrhage. In notch3 heterozygotes, we found elevated expression of both notch3 itself and target genes, indicating that specific alterations in gene expression due to partial loss of Notch3 function might contribute to the abnormalities observed in heterozygous larvae and adults. Our analysis of zebrafish notch3 mutants indicates that Notch3 regulates OPC development and mbp gene expression in larvae, and maintains vascular integrity in adults.

Distinct Notch signaling outputs pattern the developing arterial system

“…S4A), whereas only notch3 fh332 mutant embryos exhibited cranial hemorrhage (supplementary material Fig. S4A), as recently reported (Wang et al, 2014;Zaucker et al, 2013). We also noted increased numbers of ISV endothelial cells only in embryos deficient for notch1b (supplementary material Fig.…”

“…Although combining genetic manipulation of Notch signaling with the tp1-driven transgenes has been helpful, generation of receptor-specific sensors, similar to NIP-Cre, or Gal4 fusions used successfully in Drosophila (Struhl and Adachi, 1998), will be needed moving forward. jh15 nred) jh15 ], notch1a tp37 , deltac tit446 , dll4 j16e1 and notch3 fh332 have been described elsewhere (Clements et al, 2011;Covassin et al, 2009;Holley et al, 2002;Jülich et al, 2005;Leslie et al, 2007;Parsons et al, 2009;Zaucker et al, 2013). Lines generated in this study are described below.…”

Notch3 establishes brain vascular integrity by regulating pericyte number

“…Therefore, the small number of pericytes that develop in notch3 fh332 mutants is not sufficient to direct BBB formation, indicating that Notch3 function is part of a mechanism to promote adequate pericyte coverage of brain vessels. Notably, a recent study using independently identified notch3 hypomorphic alleles suggested that reduced Notch3 signaling compromised vascular tone and vascular integrity in the fins of adult zebrafish (Zaucker et al, 2013). Mural cells could not be unambiguously identified in the fin vasculature of notch3 mutants, supporting our own results.…”

Subcortical ischemic vascular disease: Roles of oligodendrocyte function in experimental models of subcortical white-matter injury

“…43 Another study showed that Notch3 regulates OPC development and myelin basic protein gene and maintains vascular integrity using zebrafish notch3 mutants. 44 Notch3 may have corresponding roles in OPC development and in encouraging vascular integrity, and also, there is a possibility that notch3 might comprise the communication between OPC and vascular cell. 44 …”

“…44 Notch3 may have corresponding roles in OPC development and in encouraging vascular integrity, and also, there is a possibility that notch3 might comprise the communication between OPC and vascular cell. 44 …”