Accumulation of Notch3 extracellular domain and not aberrant NOTCH3 signaling is the key driver of CADASIL arterial pathology
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ABSTRACT:
PROVIDER: PRJNA1014747 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR25993900.fastq.gz | Fastqsanger.gz | |||
| SRR25993901.fastq.gz | Fastqsanger.gz | |||
| SRR25993902.fastq.gz | Fastqsanger.gz | |||
| SRR25993903.fastq.gz | Fastqsanger.gz | |||
| SRR25993904.fastq.gz | Fastqsanger.gz |
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Project description:Loss of arterial smooth muscle cells (SMCs) and abnormal accumulation of the extracellular domain of the NOTCH3 (Notch3ECD) receptor are the two core features of CADASIL, the most common genetic cerebral small vessel disease. Although CADASIL is known to be caused by highly stereotyped dominant mutations in NOTCH3, the relationship between NOTCH3 receptor activity, Notch3ECD accumulation and arterial SMC loss has remained elusive, hampering the development of disease-modifying therapies. Using novel histopathological and multiscale imaging modalities, we quantified previously undetectable CADASIL-driven focal arterial SMC loss in the central nervous system of mice expressing the archetypal Arg169Cys CADASIL mutation. Notably, we found more severe arterial pathology and greater Notch3ECD accumulation in transgenic model mice overexpressing the mutation on a homozygous wild-type Notch3 background (TgNotch3R169C) than in knock-in model Notch3R170C/R170C mice expressing this mutation without a wild-type Notch3 copy. We further showed that wild-type Notch3ECD co-aggregated with mutant Notch3ECD and that elimination of one copy of wild-type Notch3 in TgNotch3R169C mice was sufficient to attenuate Notch3ECD accumulation and arterial pathology. Importantly, RNA sequencing revealed no substantial change in the expression of Notch3-regulated genes in TgNotch3R169C brain arteries. Collectively, our results provide compelling evidence that Notch3ECD accumulation, involving mutant and wild-type NOTCH3, and not aberrant NOTCH3 signaling, is the key driver of arterial SMC loss in CADASIL, thus identifying NOTCH3-lowering approaches as candidate therapeutic strategies.
2024-04-17 | GSE242794 | GEO
Project description:While most cases of vascular dementia represent complex interactions between host genetics and environmental factors, mendelian forms of vascular dementia also exist. Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), is a mendelian disease characterized by progressive vascular deterioration, cognitive deficits, and strokes. Mutations in the NOTCH3 receptor underlies the pathologies in CADASIL. NOTCH3 is primarily expressed in vascular smooth muscle cell (vSMC) and its’ expression is critical for differentiation and functional integrity of arterial vSMCs, albeit through unclear mechanism(s). To elucidate the contribution of NOTCH3 in the maintenance of cerebral vascular architecture and function, we performed micro-computed tomography (micro-CT) on the brains of aged Notch3-deficient animals. Micro-CT assessment of the cerebral vasculature architecture showed significant abnormalities including severe vessel dilation and tortuosity (dolicoectasia) of the middle cerebral artery and its branches in the Notch3-/- compared to aged-match controls. To identify the molecular pathway from NOTCH3 dysregulation to the observed cerebral vascular dysfunction, we performed single-cell RNASeq on cerebral arteries isolated from young (4w) and old (104w) Notch3-/- animals. Evaluation of the vSMC-specific transcriptomes indicated significant loss of proteins associated with muscle contraction and increased extracellular matrix production in animals that lack NOTCH3. Using a combination of immunofluorescence microscopy and in vitro functional assays, we confirmed that continued expression of Notch3 is a critical requirement for maintenance of vSMC contractile function. Impaired contractility also affected flow of cerebrospinal fluid in the parenchyma of Notch3-/- . MRI and behavioral assessments were performed in the Notch3-/- animals to elucidate the relationship between impaired vascular contractility to cognitive function. Taken together these findings link the molecular dysfunction of NOTCH3 through its regulation of vascular contractility and cerebral vessel architecture to altered neurological function and clarify the molecular pathways to cellular pathology of Notch3 driven dementias.
2024-01-17 | GSE204803 | GEO
Project description:Molecular mechanisms underlying the differentiation of brain mural cells from neural crest are poorly understood. We found that activation of Notch3 signaling in human pluripotent stem cell-derived neural crest (using lentiviral overexpression of the human Notch3 intracellular domain, N3ICD) was sufficient to direct the differentiation of mural cells. We used Notch3 ChIP-sequencing to identify cis-regulatory elements directly bound by the Notch3 transcriptional activation complex in this system.
2024-02-02 | GSE205212 | GEO
Project description:Migratory embryonal neuroblasts give rise to several tissues, including the sympathetic nervous system (SNS). Neuroblastomas are paediatric tumours of the peripheral SNS with a highly variable prognosis. We observed that high NOTCH3 expression in neuroblastomas correlated with a poor prognosis. Expression of a NOTCH3 transgene in neuroblastoma cells induced many motility genes and conferred a highly motile phenotype. Expression of these motility genes strongly correlated with NOTCH3 expression in neuroblastomas and many other tumours, suggesting a general role for NOTCH3 in regulation of these genes. Silencing of NOTCH3 or genes of the Notch-processing M-NM-3-secretase complex induced apoptosis in all neuroblastoma cell lines tested. These data suggest that NOTCH3 is a key-regulator of motility, and indispensable for survival of neuroblastoma cells. Time-course experiments of NOTCH3IC induction in two independent clones (c6 and c8), with 9 or 8 time points, respectively.
2013-08-06 | E-GEOD-16477 | biostudies-arrayexpress
Project description:Cancer immunotherapy has revolutionized the treatment of lung adenocarcinoma (LUAD); however, a significant fraction of patients fails to respond. Recent studies to understand transcriptomic determinants of immunotherapy response have pinpointed stroma-mediated resistance mechanisms. To gain a better understanding of the stromal biology at the cellular and molecular levels in LUAD, we obtained the transcriptomes of 256,379 single cells with 13,857 mesenchymal cells from 9 treatment-naïve patients. Among the mesenchymal cells, we identify subsets of FAP+PDPN+ cancer-associated fibroblasts (CAFs) and ACTA2+MCAM+ pericytes, which enrich in tumors, differentiate from lung resident fibroblasts and possess a myofibroblast phenotype. By utilizing imaging-mass cytometry, we found that both subsets are topographically adjacent to the perivascular niche and have close spatial interaction with endothelial cells (ECs). Modelling of ligand and receptor interactomes between mesenchymal and ECs identifies that NOTCH3/NOTCH signaling drives the cell-cell interaction in tumors, with pericytes and CAFs as the signal receivers and arterial and PLVAPhigh immature neovascular ECs as the signal senders. Either pharmacologically blocking NOTCH signaling or tuning down NOTCH3 level in mesenchymal cells reduced collagen production and suppressed cell invasion. By evaluating the clinical relevance of NOTCH signaling using bulk-RNA sequencing data, we demonstrate that NOTCH3 correlates with poor survival in stroma-rich patients and that a T-cell inflammation gene signature predicts survival only in low-NOTCH3 expressing patients. Our study underscores the importance of NOTCH3 in regulating tumor stroma biology and may serve as a potential target for immunotherapy combination strategies.
2024-01-31 | GSE253013 | GEO
Project description:Molecular defects in some ultra-rare subtypes of familial lipodystrophies remain unidentified. We identified novel NOTCH3 heterozygous variants in familial partial lipodystrophy (FPL) pedigrees. All variants were clustered in the heterodimerization domain of the negative regulatory region of NOTCH3. Proteomics of skin fibroblasts revealed significantly higher RNA expression of NOTCH3 and activation of widespread senescence pathways in the FPL patients versus controls.
2024-10-28 | MSV000096235 | MassIVE
Project description:Cerebral small vessel disease (SVD) is a prevalent disease of aging and a major contributor to stroke and dementia. The most commonly inherited SVD, CADASIL, is caused by dominantly acting cysteine-altering mutations in NOTCH3. These mutations change the number of cysteines from an even to an odd number, but the impact of these alterations on NOTCH3 protein structure remain unclear. Here, we prepared wildtype and four mutant recombinant NOTCH3 protein fragments to analyze the impact of CADASIL mutations on oligomerization, thiol status, and protein stability. Using gel electrophoresis, tandem MS/MS, and collision-induced unfolding, we find that NOTCH3 mutant proteins feature increased amounts of inappropriate disulfide bridges, reduced cysteines, and structural instability. Presence of a second protein factor, an N-terminal fragment of NOTCH3 (NTF), is capable of further altering disulfide statuses of both wildtype and mutant proteins, leading to increased numbers of reduced cysteines and further destabilization of NOTCH3 structure. In sum, these studies identify specific cysteine residues alterations and quaternary structure induced by CADASIL mutations in NOTCH3; further, we validate that reductive factors alter the structure and stability of this small vessel disease protein.
2022-05-20 | PXD031097 | Pride
Project description:Vascular smooth muscle cells (VSMC) are important for contraction, blood flow distribution and regulation of blood vessel diameter, but to what extent they contribute to the integrity of blood vessels and blood-brain barrier function is less well understood. In this report, we explored the impact of the progressive loss of VSMC in the Notch3-/- mouse on blood vessel integrity in the central nervous system To explore the molecular consequences of the VSMC phenotype in Notch3-/- mice, we performed genome-wide transcriptional profiling of the brain vasculature in mutants and littermate controls using Affymetrix Mouse Genome 430 2.0 Array. Three Notch3 knockout and three littermate control mice at age of 2 months were used to profile the transcriptomes. Total RNA was extracted from cerebral and cerebellar microvascular fragments and hyrbridized separately on the Mouse Genome 430 2.0 Array according to standard procedures.
2015-02-18 | E-GEOD-55203 | biostudies-arrayexpress
Project description:Notch is a major oncogenic driver in T cell acute lymphoblastic leukemia (T-ALL), in part because it binds to an enhancer that increases expression of MYC. Here, we exploit the capacity of activated NOTCH1 and NOTCH3 to induce T-ALL, despite substantial divergence in their intracellular regions, as a means to elucidate a broad, common Notch-dependent oncogenomic program through systematic comparison of the transcriptomes and Notch-bound genomic regulatory elements of NOTCH1- and NOTCH3-dependent T-ALL cells. ChIP-seq studies show a high concordance of functional NOTCH1 and NOTCH3 genomic binding sites that are enriched in binding motifs for RBPJ, the transcription factor that recruits activated Notch to DNA. The interchangeability of NOTCH1 and NOTCH3 was confirmed by rescue of NOTCH1-dependent T-ALL cells with activated NOTCH3 and vice versa. Despite remarkable overall similarity, there are nuanced differences in chromatin landscapes near critical common Notch target genes, most notably at a Notch-dependent enhancer that regulates MYC, which correlates with responsiveness to Notch pathway inhibitors. Overall, a common oncogenomic program driven by binding of either Notch is sufficient to maintain T-ALL cell growth, whereas cell-context specific differences appear to influence the response of T-ALL cells to Notch inhibition.
2017-09-27 | GSE104261 | GEO