We performed a survey of sequence variation in a series of 20 genes involved in inflammation-related pathways for association with dementia risk in twin and unrelated case-control samples consisting in total of 1462 Swedish dementia casesand 1929 controls. For a total of 218 tested genetic markers,
strong evidence was obtained implicating a region near AGER and NOTCH4 on chromosome 6p with replication across both samples and maximum combined significance at marker rs1800625 (OR = 1.37, 95% CI 1.19-1.56, p = 1.36x10(-6)). Imputation of the associated genomic interval provided an improved signal atrs8365, near the 3UTR of AGER (p = 7.34x10(-7)). The associated region extends 120 kb encompassing 11 candidate genes.While AGER encodes a key receptor for amyloid-beta protein, an analysis of network context based upon genes now confirmed to contribute to dementia risk (AbetaPP, PSEN1, PSEN2, CR1, CLU, PICALM, and APOE) suggested strong functional coupling to NOTCH4, with no significant coupling to the remaining candidates. The implicated region occurs in the broad HLA locus on chromosome 6p, but associated markers were not in strong LD with known variants that regulate HLA gene function, suggesting that this may represent a signal distinct from immune-system pathways.
Suzuki T, etal., Int J Oncol. 2000 Dec;17(6):1131-9.
The development of cancer is a cellular process that reflects and is partly driven by alterations in cell determination. Mutations in various genes responsible for cell determination have been identified as being oncogenic; however, little is known about the involvement of normal cell fate-determini
ng mechanisms in the oncogenic process. The Notch pathway is an evolutionarily conserved, general cell-interaction mechanism that controls fundamental aspects of cell determination during the development of vertebrates and invertebrates. We have explored the roles of the recently cloned human Notch4 gene, which is a homologue of Drosophila Notch, in endometrial tissue, a cellular environment where cell fate changes take place and neoplastic conditions have been characterized. Northern blot analysis, dot blot analysis, and in situ hybridization were performed on samples of normal endometria at different phases of the menstrual cycle and endometrial cancers. Our results showed that: a) Both human Notch4 and translocation-associated Notch homologue-1 (TAN-1), another human homologue of Drosophila Notch, are expressed in the normal endometrium, mainly in the endometrial glandular cells. b) The level of expression of human Notch4, but not TAN-1, changes during the normal menstrual cycle; i.e., the level of Notch4 expression was significantly lower during the secretory phase than during the proliferative phase. c) Endometrial cancers express a significantly lower level of human Notch4 and a significantly higher level of TAN-1 than normal endometria. These results suggest that human Notch4 and/or TAN-1 are involved in the changes of the endometrium during the menstrual cycle, and also in the development of endometrial cancer.
Luo X, etal., Biol Psychiatry 2004 Jan 15;55(2):112-7.
BACKGROUND: The goal of this study was to investigate the relationship between the NOTCH4 gene and schizophrenia in African American (AA) and European American (EA) subjects. METHODS: Two single nucleotide polymorphisms (SNPs) at the NOTCH4
;'>NOTCH4 locus were genotyped in 123 AA schizophrenia patients, 223 EA schizophrenia patients, 85 AA healthy control subjects, and 211 EA healthy control subjects. The specific markers studied were -1725T/G and -25T/C. Comparisons of allele and haplotype frequencies between patients and control subjects were performed with the chi-square test, the Fisher's Exact Test, and CLUMP software. Linkage disequilibrium (LD) between these two SNPs was calculated with the 3LOCUS program. RESULTS: The haplotype -1725G/-25T associates to schizophrenia in AA subjects (p =.0008), but not in EA subjects. Alleles -1725G and allele -25T are in positive LD both in AAs and EAs. Allele and haplotype frequencies differ significantly between AAs and EAs. CONCLUSIONS: The haplotype -1725G/-25T at the NOTCH4 locus, which results from SNPs of NOTCH4 that are in LD, may increase susceptibility to schizophrenia in AAs. Any effect of this locus on risk for schizophrenia is population-specific.
Uyttendaele H, etal., Microvasc Res 2000 Sep;60(2):91-103.
The mouse Notch4 gene is expressed specifically in endothelial cells. Notch4/int-3, a truncated form of Notch4, acts as a constitutive activated Notch receptor. We used rat brain microve
ssel endothelial cells (RBE4) to study the role of Notch4 and Jagged-1 in endothelial cell differentiation. Both Notch4/int-3 and Jagged-1 were able to induce microvessel-like structures with morphological and biochemical properties similar to brain endothelial microvessels. Ectopic expression of full-length Notch4 did not effect RBE4 cells. Activation of the Notch signal transduction pathway was measured by the induction of endogenous Notch4 and Jagged-1 genes and of Jagged-1 proteins. The observed morphological changes to RBE4 cells correlated with endogenous Notch4 and Jagged-1 gene activation. Our observations demonstrate that Notch signaling can promote endothelial cell differentiation and morphogenesis.
Wang Z, etal., Am J Med Genet B Neuropsychiatr Genet. 2006 Dec 5;141B(8):902-6.
This work reviewed all the reports on the NOTCH4 gene in schizophrenia, which have been published since the gene was found to be associated with illness among a British population in 2000. The results from independent studies were inconsistent. Allelic heteroge
neity, clinical diagnosis, ethnical backgrounds, and linkage disequilibrium (LD) structures in the human genome may be major reasons for poor replication. A couple of studies suggested that the NOTCH4 gene could play a role in a subgroup of the disease, such as early-onset schizophrenia and negative symptoms. A single study revealed a weak association of the NOTCH4 gene with frontal lobe brain volumes and a strong association with frontal lobe cognitive performance. A meta-analysis showed stronger evidence of the NOTCH4 association in family-based studies than in case-control studies. In a previous study, we found that rs520692, a single nucleotide polymorphism (SNP) at the NOTCH4 locus, was associated with schizophrenia in a Chinese population. In the present study, we applied a large sample size to re-evaluate our initial findings and then confirmed the rs520692 association with illness. The pairwise measures did not show strong LD between paired SNPs although the SNPs tested are located within a 34-kb region, suggesting that LD within the NOTCH4 gene has been broken rapidly by historical recombination in the Chinese population. Taken together, the NOTCH4 gene may be associated with schizophrenia but how the gene contributes to the etiology of the illness needs a further investigation.
Tochigi M, etal., Am J Med Genet B Neuropsychiatr Genet. 2004 Jul 1;128B(1):37-40.
The NOTCH4 gene is located at 6p21.3 and involved in the development and patterning of the central nervous systems. Recently, Wei and Hemmings [2000] observed that the gene was associated with schizophrenia. Subsequent to the report, several studies investigated
the gene in schizophrenia, with controversial and inconclusive results. In the present study, we investigated six polymorphisms (SNPs 1-5 and a CTG repeat) of the gene in Japanese subjects with schizophrenia (n = 284) and the same number of controls. The polymorphisms include SNP5, which has been observed to be associated with schizophrenia in a Chinese population and two new SNPs 3-4 adjacent to SNP5, in addition to the SNPs 1-2 and the CTG repeat, which were suggested for the association with the disease in the previous study. As a result, no significant difference in genotypic distributions or allelic frequencies of the six polymorphisms of the gene was observed between the patients and the controls. Also, no significant difference was found in frequencies of haplotypes of the six polymorphisms between the patients and the controls. However, the distribution of SNP2 was significantly deviated from Hardy-Weinberg equilibrium in the patients (P = 0.000986), not in the controls, which could be a chance or due to an association of SNP2 with the disease. In conclusion, the present study provided no clear evidence for an association between the NOTCH4 gene and schizophrenia in the Japanese population.
Breast cancers (BCs) typically express estrogen receptors (ERs) but frequently exhibit de novo or acquired resistance to hormonal therapies. Here, we show that short-term treatment with the anti-estrogens tamoxifen or fulvestrant decrease cell proliferation but increase BC stem cell (BCSC) activity
through JAG1-NOTCH4 receptor activation both in patient-derived samples and xenograft (PDX) tumors. In support of this mechanism, we demonstrate that high ALDH1 predicts resistance in women treated with tamoxifen and that a NOTCH4/HES/HEY gene signature predicts for a poor response/prognosis in 2 ER+ patient cohorts. Targeting of NOTCH4 reverses the increase in Notch and BCSC activity induced by anti-estrogens. Importantly, in PDX tumors with acquired tamoxifen resistance, NOTCH4 inhibition reduced BCSC activity. Thus, we establish that BCSC and NOTCH4 activities predict both de novo and acquired tamoxifen resistance and that combining endocrine therapy with targeting JAG1-NOTCH4 overcomes resistance in human breast cancers.
Zhang B, etal., Biomed Res Int. 2015;2015:408096. doi: 10.1155/2015/408096. Epub 2015 Oct 28.
NOTCH4 regulates signaling pathways associated with neuronal maturation, a process involved in the development and patterning of the central nervous system. The NOTCH4 gene has also been identified as a possible susceptibili
ty gene for schizophrenia (SCZ). The objective of this study was to examine the relationship between NOTCH4 polymorphisms and SCZ in the Chinese Han population. The rs2071287 and rs204993 polymorphisms of the NOTCH4 gene were analyzed in 443 patients with SCZ and 628 controls of Han Chinese descent. Single SNP allele-, genotype-, and gender-specific associations were analyzed using different models (i.e., additive, dominant, and recessive models). This association study revealed that the rs204993 polymorphism is significantly associated with susceptibility for SCZ and that the AA genotype of rs204993 is associated with a higher risk for SCZ (P = 0.027; OR = 1.460; 95% CI, 1.043-2.054). Our data are consistent with those obtained in previous studies that suggested that rs204993 is associated with SCZ and that the AA genotype of rs204993 demonstrates a higher risk. Further large-scale association analyses in Han Chinese populations are warranted.
Shibata N, etal., J Gerontol A Biol Sci Med Sci. 2007 Apr;62(4):350-1.
It is shown that Notch 4 plays important roles in the pathogenesis of Alzheimer's disease (AD). To investigate whether three single nucleotide polymorphisms (SNPs) of the Notch4 gene are associated with AD, the three SNPs were genotyped by a polymerase chain rea
ction-restriction fragment length polymorphism method for 243 AD patients and 130 age-matched controls. We also confirmed the linkage disequilibrium among these three SNPs of the gene using the EH program. The three SNPs did not seem to alter risk for AD. Our study suggests that SNPs studied are not associated with AD. The linkage disequilibrium of this locus indicates that there is genetic heterogeneity in the Notch4 gene. We could not confirm the previous synergetic associations of the 5' untranslated region (rs367398) C/C genotype in apolipoprotein E epsilon4 bearers in AD patients. Potential markers nearby the 5' untranslated region polymorphism might affect risk for AD.
Gao J, etal., Int J Oncol. 2016 Jan;48(1):329-37. doi: 10.3892/ijo.2015.3221. Epub 2015 Oct 30.
Deregulated expression of Notch receptors and abnormal activity of Notch signaling have been observed in a growing number of malignant tumors, however, the expression and activity of Notch in hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC) and their relationship with HBV X protein
(HBx) are still not fully elucidated. To address this, we examined the overall expression of Notch receptors in HBV-associated HCC tissues, analyzed their relationship with HBx, and further investigated the role of Notch signaling in HBx stable transfected HepG2 cells (HepG2X). The results showed that Notch signaling could be activated by HBx in HepG2 cells. The expression of cytoplasmic Notch1 or nuclear Notch4 was correlated with the expression of HBx in HBV-associated HCC tissues. The expression of cytoplasmic Notch1 or nuclear Notch4 could also be upregulated by HBx in HepG2X cells. The upregulation of Notch1 by HBx was through p38 MAPK pathway. Moreover, HBx was found to directly interact with Notch1, whereas, not with Notch4 in HepG2X cells. Suppression of Notch signaling by gamma-secretase inhibitor (GSI) decreased cell growth, blocked cell cycle progression and induced cell apoptosis in HepG2X cells. The present study indicates that HBx activates Notch signaling by its effects on Notch1 and Notch4, and therefore, recruits Notch signaling as a downstream pathway contributing to its carcinogenic role in HBV-associated HCC.
OBJECTIVE: Notch signaling pathway controls key functions in vascular and endothelial cells (EC). However, little is known about the role of Notch in allografted vessels during the development of transplant arteriosclerosis (TA). This study investigated regulation of the Notch pathway on cardiac all
ograft arteriosclerosis and further examined its implication in EC dysfunction. METHODS AND RESULTS: Here we show that, among Notch receptors, Notch2, -3, and -4 transcript levels were markedly downregulated in TA compared to tolerant and syngeneic allografts. TA correlates with high levels of tumor necrosis factor (TNF), transforming growth factor (TGF)beta, and IL10, which consistently decrease Notch4 expression in transplants and cultured ECs. We found that inhibition of Notch activity, reflected by both a reduced CBF1 activity and Hes1 expression, parallels the downregulation of Notch4 expression mediated by TNF in ECs. Notch4 and Hes1 knockdown enhances vascular cell adhesion molecule-1 expression and promotes EC apoptosis. Silencing Notch4 or Hes1 also drastically inhibits repair of endothelial injury. Overall, our results suggest that Notch4 and basal Notch activity are required to maintain EC quiescence and for optimal survival and repair in response to injury. CONCLUSIONS: Together, our findings indicate that impaired Notch4 activity in graft ECs is a key event associated with TA by triggering EC activation and apoptosis.
Raafat A, etal., Oncogene. 2004 Dec 16;23(58):9401-7.
Recently, we have identified a novel 1.8 kb human Notch4/Int3 RNA species (designated h-Int3sh). The h-Int3sh RNA encodes a protein that is missing the CBF1-binding region (RAM23) of the Notch 4/Int3 intracellular domain (ICD). Expression of h-Int3sh in the MCF1
0A 'normal' human mammary epithelial cell line has been previously shown to induce changes characteristic of oncogenic transformation, including anchorage-independent growth in soft agar. To study the consequences of h-Int3sh expression in vivo on mammary gland development and tumorigenesis, three transgenic mouse lines were established, in which the transgene is the Whey acidic protein (WAP) promoter linked to h-Int3sh. Expression of WAP-Int3sh was detectable in the mammary gland at day 15 of pregnancy in each transgenic line. Mammary gland development in all founder lines is normal and the females can lactate. WAP-h-Int3sh females from each of the founder lines develop mammary tumors, but with a long latency (average age of 18 months). Tumor development was associated with activation of Notch pathway, as evidenced by upregulation of Hes-1. The long latency of mammary tumors in WAP-h-Int3sh mice could be due in part to the subcellular localization of h-Int3sh. Immunofluorescence analysis of transfected COS-1 cells showed that h-Int3sh is localized in the cytoplasm and nucleus, while Int3-ICD is detected only in the nucleus. We speculate that the Notch4/Int3 ICD-induced block to mammary gland development and tumorigenesis are consequences of an increasing gradient of CBF1-dependent Notch4/Int3 signaling.
Alopecia areata (AA) is a disorder primarily affecting the hair and nails in which associated autoimmune or atopic disease is common. Genetically, it is a complex trait with evidence of a role for genes of the major histocompatibility complex (MHC), the interleukin-1 cluster and chromosome 21 in the
pathogenesis. The strongest association is with HLA class II alleles, although whether this indicates a direct contribution to the pathogenesis or results merely from linkage disequilibrium with nearby disease genes is unknown. Notch4 is a recently defined gene in the HLA class III region. Notch signalling is a direct determinant of keratinocyte growth arrest and entry into differentiation. A possible role for Notch in hair growth has been indicated by transgenic mouse findings that activation of the Notch pathway in the hair cortex leads to aberrant differentiation of adjacent hair-shaft layers. Notch4 is therefore a plausible candidate gene for AA. We have examined two polymorphisms in the coding sequence of the Notch4 gene at positions +1297 and +3063 in a case-control study of 116 AA patients and 142 ethnically matched, healthy control subjects. The initial analysis showed a significant association of AA in the overall data set with the Notch4(T+1297C) polymorphism (P<0.001) but not with Notch4(A+3063G). To confirm this association, we genotyped an additional 62 patients and found that the risk for disease was higher in Notch4(+1297C) homozygotes [odds ratio (OR) 3.43 (1.63, 7.19)] than in heterozygotes [OR 2.58 (1.57, 4.24)]. On classifying the patients by severity of disease, the association appeared to be confined to the severest form (alopecia universalis) [OR 4.02 (1.64, 9.88), P=0.0014]. These results support previous findings showing that different HLA susceptibility alleles are associated with mild and severe AA.
Verhein KC, etal., Environ Health Perspect. 2015 Aug;123(8):799-805. doi: 10.1289/ehp.1408852. Epub 2015 Feb 6.
BACKGROUND: Ozone is a highly toxic air pollutant and global health concern. Mechanisms of genetic susceptibility to ozone-induced lung inflammation are not completely understood. We hypothesized that Notch3 and Notch4 are important determinants of susceptibilit
y to ozone-induced lung inflammation. METHODS: Wild-type (WT), Notch3 (Notch3-/-), and Notch4 (Notch4-/-) knockout mice were exposed to ozone (0.3 ppm) or filtered air for 6-72 hr. RESULTS: Relative to air-exposed controls, ozone increased bronchoalveolar lavage fluid (BALF) protein, a marker of lung permeability, in all genotypes, but significantly greater concentrations were found in Notch4-/- compared with WT and Notch3-/- mice. Significantly greater mean numbers of BALF neutrophils were found in Notch3-/- and Notch4-/- mice compared with WT mice after ozone exposure. Expression of whole lung Tnf was significantly increased after ozone in Notch3-/- and Notch4-/- mice, and was significantly greater in Notch3-/- compared with WT mice. Statistical analyses of the transcriptome identified differentially expressed gene networks between WT and knockout mice basally and after ozone, and included Trim30, a member of the inflammasome pathway, and Traf6, an inflammatory signaling member. CONCLUSIONS: These novel findings are consistent with Notch3 and Notch4 as susceptibility genes for ozone-induced lung injury, and suggest that Notch receptors protect against innate immune inflammation.
