NOTCH4 Locus Research Articles

Association of the NOTCH4 gene polymorphism with schizophrenia in the Indian population

Schizophrenia is a chronic serious neuropsychiatric disorder with a prevalence of 1% worldwide. Neurogenic locus NOTCH homolog protein 4 known as the NOTCH4 gene is located on chromosome 6p. NOTCH4 protein is a transmembrane protein that plays a major role in cell signaling pathways and cell differentiation. NOTCH4 SNP rs367398 is located in the gene's largest intron, intron 18, which is an area of known importance in transcriptional regulation and may be related to schizophrenia. Although many studies have shown the association of NOTCH4 polymorphism with schizophrenia, the exact molecular mechanism behind this still unknown. This study was aimed to determine the frequency of NOTCH4 rs367398 & rs387071 polymorphism and its association with schizophrenia. In total, 150 schizophrenia patients and 100 healthy controls were recruited from psychiatry department after obtaining written informed consent. NOTCH4 polymorphism was genotyped using PCR-RFLP. Positive and Negative Syndrome Scale (PANSS) and Global assessment of functioning (GAF) were used to assess the severity of schizophrenia. Both the NOTCH4 rs367398 & rs387071 genotypes were significantly associated with schizophrenia. On gender based analysis, the rs387071 was significantly associated with schizophrenia in males and whereas the rs367398 is associated with females of the study group. Further, the haplotype analysis has also supported the association of NOTCH4 locus with schizophrenia (Global haplotype association p = 0.021). The findings of the study suggest that the NOTCH4 polymorphism is significantly associated with schizophrenia in Indian population. These SNPs might also have an important role in the symptomatology of the disease.

Large‐scale structural variations linked to the NOTCH4 locus of the Human Major Histocompatibility Complex

The human major histocompability complex (MHC) encodes antigen‐presenting proteins and other proteins critical to the function of the the immune system. It is also the most polymorphic component of the human genome. Variations within the MHC region have been linked to both susceptibility and resistance to multiple autoimmune diseases. The objective of the present study is to determine whether the portion of the MHC linked to the NOTCH4 gene is characterized by large‐scale structural variation and if it is, whether that variation is linked to auto‐immune disease. The NCBI human genome reference sequence indicates that NOTCH4 is contained within a 450 kilobase PmeI restriction fragment. To date, this study has used polymerase chain reaction to isolate NOTCH4 specific probes from cDNA for NOTCH4. The probes will be hybridized with Southern blots of pulsed field electrophoresis gels of PmeI digested genomic DNA isolated from patients with systemic lupus erythematosus and normal controls. This will allow the observation of size variation and imply the possibility of copy number variations, insertions and/or deletion within the region and their possible role in autoimmune disease.

Search for schizophrenia susceptibility variants at the HLA-DRB1 locus among a British population

Schizophrenia is a complex mental disorder with unknown aetiology. Both candidate gene and genome-wide association (GWA) studies suggest that the human leukocyte antigen (HLA) system may play a part in development of the illness, but the causal HLA variant(s) remain(s) unclear. Previous studies showed that the DRB1*0101 and DRB1*13 alleles might be associated with a high risk of schizophrenia. Therefore, the present study was undertaken to test their association with the disease by genotyping seven DRB1-tagging single nucleotide polymorphisms (SNPs) in a British population. The results showed that, of the previously reported variants that were associated with schizophrenia, the DRB1*1303 allele was the only one marginally associated with a protective effect on the illness in our sample set (χ² = 4.138, P = 0.042, odds ratio (OR) = 0.42, 95 % confidence interval (CI) 0.27-0.66). Interestingly, a significant association was found for rs424232 (χ² = 9.404, P = 0.002, OR = 0.69, 95 % CI 0.54-0.88), which is a tag SNP for the DRB1*1303 allele and located near to the NOTCH4 gene that is a schizophrenia susceptibility locus confirmed by GWA studies. Analysis with the Haploview program demonstrated that rs424232 was in complete linkage disequilibrium with rs3130297 and rs3131296 present in the NOTCH4 locus. While we have failed to confirm association of the candidate alleles in the DRB1 gene with a high risk of schizophrenia, the present work suggests that the association signal detected in the HLA class II locus may extend a relatively long distance, and more work is needed in order to identify the true causal variants within this region or nearby.

Mouse mammary tumor virus derived from wild mice does not target Notch-4 protooncogene for the development of mammary tumors in inbred mice

The colony of wild mice, named Jyg, has been shown to express an exogenous mouse mammary tumor virus (Jyg-MMTV). This virus induces mammary tumors in its natural host at a high incidence (≈ 80%) resulting from insertion mutations in Notch-4 (43%), Wnt-1 (26%), and Fgf-3 (13%). Since the activation of Notch-4 is not common in mammary tumors of standard laboratory strains of mice infected with various MMTV strains, we examined the consequences of Jyg-MMTV infection in BALB/c and C57BL/6 mice. The results show that Jyg-MMTV induces mammary tumors in both mouse strains, but the incidence of mammary tumors in BALB/c mice is greater than in C57BL/6 mice. Surprisingly, however, none of the 75 mammary tumors, analyzed both by Southern and Northern hybridizations, showed insertion mutations in or expression of Notch-4. In contrast, both Wnt-1 and Fgf-3 were found to be involved in these tumors. Our findings may suggest, among other possibilities, the existence of a structural difference(s) between laboratory and wild mice at the Notch-4 locus that regulates the integration of Jyg-MMTV proviral DNA.

Association study between the TNXB locus and schizophrenia in a Japanese population

The chromosome 6p21-24 region, which contains the human leukocyte antigen (HLA) region, has been suggested as an important locus for a susceptibility gene for schizophrenia. Recently, a significant association between schizophrenia and the TNXB locus, located immediately telomeric of the NOTCH4 locus in the HLA region, was observed. Few studies have further investigated the region in schizophrenia. In the present study, we investigated the region in a Japanese population. Subjects included 241 patients with schizophrenia and 290 controls. Twenty-six single nucleotide polymorphisms (SNPs) and the corresponding haplotypes were analyzed. As a result, exactly the same SNPs in the TNXB locus (rs1009382 and rs204887) as in the previous study were associated with schizophrenia (P = 0.034 and 0.034, respectively, uncorrected). A SNP (rs2071287) in the NOTCH4 locus and haplotype around it were also suggested to associate with the disease, consistent with another previous study (P = 0.041 and permutation P = 0.024, respectively, uncorrected). Although these associations became insignificant after Bonferroni correction, the findings might provide support for the association of the TNXB locus or its adjacent region of the NOTCH4 locus with schizophrenia.

A review and re‐evaluation of an association between the NOTCH4 locus and schizophrenia

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 heterogeneity, 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.

Notch 4 Plymorphism Associated with Risk of Aplastic Anemia.

Aplastic anemia (AA) is a heterogeneous disease characterized by bone marrow aplasia and peripheral blood pancytopenia, a profound deficit of hematopoietic stem and progenitor cells. Although the causes for aplastic anemia are multiple, the pathogenic mechanism leading to the disease might be limited to two main pathways: environmental factors and genetic susceptibility. The mammalian Notch family has four homologs, Notch1 to 4. Notch4 signaling plays a role in lineage commitment in hematopoiesis. The human Notch4 gene is located at the MHC on chromosome 6p21.3. Many single nucleotide polymorphisms (SNP) have been mapped to the Notch4 locus. Therefore, we hypothesized that some of these variants could potentially influence the expression level of Notch4 protein. To investigate the role of the two Notch4 rs2071282 (+2763 C>T, Pro204Leu) and 422951(+3323 A>G, Thr320Ala) polymorphisms in susceptibility to AA, a case-control study was conducted in Chonnam National University Hwasun Hospital between March 2002 and June 2006. We genotyped in 151 AA patients and 552 healthy control subjects using PCR-RFLP. Here, we report that rs2071282 CT genotype is significantly associated with increased risk for AA. Using subjects with the rs2071282 CC as a reference group, the odds ratio (OR) of CT is 1.77 (95% CI 1.05–2.98, p=0.032). The haplotype CG is significantly associated with the risk of AA. Using subjects with the haplotype CA, the OR of haplotype CG is 0.70 (95% CI 0.50–0.98, p= 0.037).In conclusion, we were able to identify two polymorphisms of Notch4, therefore, theses results suggest that Notch polymorphism may play an important role in the susceptibility to AA.Distribution of Notch polymophisms in aplastic anemia patients and controlsControl (%)AA(%)OR (95% CI)pRs 422951AA302(54.8)94(62.3)1AG216(39.2)49(32.5)0.72(0.478–1.098)0.128GG33(6.0)8(5.3)0.54(0.23–1.279)0.163Total551151Rs 2071282CC481(87.1)120(81.1)1CT68(12.3)28(18.9)1.769(1.049–2.982)0.032TT3(0.5)0--TotalHaplotype frequency for AA and controlsHaplotypeControl (%)AA(%)P (Chi-square)OR (95% CI)pC-A743(67.7)206(70.0)0.0621C-G281(25.6)60(20.4)0.70(0.50–0.98)0.037T-A72(6.5)28(9.5)1.49(0.90–2.47)0.126*Haplotype frequencies were estimated from genotype data using Gibbs- algorithm.*OR and 95%Cis were estimated using based logistc regression and adjusted for sex and age

No evidence for association between NOTCH4 and schizophrenia in a large family-based and case–control association analysis

An analysis of 80 British parent-offspring trios by Wei and Hemmings in 2000 revealed thre1e out of five markers within the NOTCH4 locus to be strongly associated with schizophrenia. In our present study, we have examined NOTCH4 markers in large samples of German and Palestinian-Arab origin. Our study population comprised a German case-control sample (n=512 schizophrenia patients and n=232 controls) and two independent parent-offspring trio samples of German (n=159 trios) and Palestinian-Arab (n=208 trios) descent. We examined a total of ten single nucleotide polymorphisms within the NOTCH4 locus and the adjacent loci, spanning a region of approximately 100 kb. Neither single marker nor haplotype analyses showed association with schizophrenia. In addition, analyses of the German case-control and trio samples revealed no significant association between NOTCH4 polymorphisms and early-onset schizophrenia. Our results suggest that NOTCH4 is unlikely to play a major role in the genetic predisposition to schizophrenia in the German or the Palestinian-Arab population.

Neither single-marker nor haplotype analyses support an association between genetic variation near NOTCH4 and bipolar disorder.

Markers near the NOTCH4 locus on chromosome 6p21.3 have been reported to be associated with schizophrenia in some studies. Since schizophrenia and bipolar affective disorder (BPAD) may share genetic determinants, we tested markers in and near NOTCH4 in a sample of 153 parent-offspring triads ascertained through a sibling pair with BPAD for evidence of association. This sample would have 80% power to detect an association at or above a genotype relative risk of 2.4 at the 10(-7) level of significance. In addition to the two markers previously showing the most significant association with schizophrenia, three additional nearby markers were studied. The five markers were genotyped using validated methods. Both single-marker and 3-marker haplotype data was analyzed using family-based association methods. No genome-wide significant association was detected between any of the five SNP-markers and BPAD in this sample. One marker showed nominal evidence of association (P = 0.049), but this evidence was not supported by haplotype analyses including nearby flanking markers or by case-control analysis using 93 Caucasian controls. These results do not support an association between genetic variation near NOTCH4 and BPAD in this sample.

Is NOTCH4 associated with schizophrenia?

The NOTCH4 locus was reported to be associated with schizophrenia in our previous study but the subsequent replication by other workers has been inconsistent. To find out possible reasons for the poor replication, the present work was undertaken to analyse four functional single nucleotide polymorphisms (SNPs) (rs367398, rs915894, rs520692 and rs422951) at the NOTCH4 locus among 141 schizophrenic family trios of Chinese Han descent. Of these four SNPs, rs520692 was the only one associated with schizophrenia (P = 0.017); the other three, however, did not show any association with the illness, including rs367398 located in the promoter region, which had shown a strong association with the illness in our previous study conducted with British samples. Although these four SNPs analysed lie within a less than 4 kb segment of genomic DNA, the pattern of linkage disequilibrium between them was unexpected. The strongest linkage disequilibrium was shown only between rs367398 and rs520692 and between rs520692 and rs422951 in both parent and patient groups. This study raises the possibility that there might be two or more disease-underlying variants at the NOTCH4 locus or at a nearby locus, and that the allelic or locus heterogeneity may be one of the possible reasons for the poor replication of the NOTCH4 finding.

Lack of a genetic association between the TNXB locus and schizophrenia in a Chinese population

A recent study demonstrated that the tenascin X (TNXB) gene was associated with schizophrenia in a British population. To replicate the initial finding, we analysed two positive single nucleotide polymorphisms (SNPs), rs1009382 and rs204887 present at the TNXB locus, in a Chinese population by using PCR-based restriction fragment length polymorphism analysis. We recruited a total of 136 family trios consisting of fathers, mothers and affected offspring with schizophrenia. The transmission disequilibrium test did not show allelic association between these two SNPs and schizophrenia, and the rs1009382-rs204887 haplotypes were not associated with the illness either. The present results suggest that the TNXB locus does not appear to be associated with schizophrenia in the Chinese population. Because the TNXB gene is less than 100 kb away from the NOTCH4 locus that was also reported to be associated with schizophrenia, allelic and locus heterogeneity could be possible reasons for the failure to replicate the TNXB finding.

NOTCH4 gene haplotype is associated with schizophrenia in African Americans

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 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.

TNXB locus may be a candidate gene predisposing to schizophrenia.

We report here on the detection of nine single nucleotide polymorphisms (SNPs) near to the NOTCH4 locus in the search for schizophrenia susceptibility genes in the class III region of the human major histocompatibility complex (MHC). We totally analyzed 122 family trios recruited in the UK. The TDT analysis demonstrated that of the nine SNPs, three were associated with schizophrenia, including rs1009382 (P = 0.00047), rs204887 (P = 0.007), and rs8283 (P = 0.015). Both rs1009382 and rs204887 are present in the TNXB locus. The rs1009382 is a non-synonymous SNP located in exon 23 of the gene and its A to G base change causes a Glu2578Gly substitution. The goodness-of-fit test showed that genotypic distribution of rs1009382 was deviated from Hardy-Weinberg equilibrium due to homozygote excess in the patient group (P = 0.01), suggesting that a double dose of a genetic risk may be involved. Possibly, rs1009382 is a candidate SNP predisposing to a schizophrenic illness. Moreover, the test for linkage disequilibrium (LD) between paired SNPs showed that the nine SNPs studied may be in the same LD block with an unexpected pattern as the strength of LD was not correlated with the distance between paired SNPs. The haplotype analysis suggested that there might be more than one disease-related allele located in the class III region of the MHC, and that these alleles possibly confer either susceptibility or resistance to schizophrenia.

Two NOTCH4 polymorphisms and their relation to schizophrenia susceptibility and different personality traits

Recently, linkage disequilibrium mapping of the major histocompatibility complex region on the short arm of human chromosome 6 suggested that the NOTCH4 locus is highly associated with schizophrenia. We analysed two polymorphisms in this gene in Swedish schizophrenic patients ( =74) and control subjects ( =135). The NOTCH4 variants were also analysed in schizophrenic patients with regard to subdiagnosis, age at first hospitalization, abuse/dependence of alcohol, solvents, or drugs, previous suicide attempts, extrapyramidal symptoms, treatment with anticholinergic drugs, and response to anti-psychotic drug treatment. Control subjects were scrutinized with regard to personality, another partially heritable trait suggested being of importance in schizophrenia. In addition, two intermediate endophenotypes suggested being of importance in schizophrenia, dopamine D(2) receptor density in striatum and monoamine metabolites in cerebrospinal fluid, respectively, were investigated with regard to the two NOTCH4 variants. There was no significant association between the patients and the controls for the two investigated polymorphisms neither for the parameters analysed in the schizophrenia material. The NOTCH4 SNP2 variant, an A-->G substitution, was associated with the Karolinska Scales of Personality Irritability scale. The NOTCH4 (CTG)(n) variant was associated with the revised NEO personality inventory Extraversion and Activity (E4) scales. However, after correction for multiple testing, no difference remained significant. The results for the endophenotypes and the polymorphisms were non-significant. The present study does not support that the investigated NOTCH4 variants have a major influence on susceptibility to schizophrenia or related neurobiological traits.

Genetic analysis of the (CTG)n NOTCH4 polymorphism in 65 multiplex bipolar pedigrees

A strong genetic association between the NOTCH4 locus on chromosome 6 and schizophrenia was recently reported. Based on the data suggesting overlapping susceptibility for schizophrenia and bipolar disorder, we genotyped the polymorphic (CTG)n encoding polyleucine repeat in exon 1 of NOTCH4 in 65 pedigrees ascertained for a genetic linkage study of bipolar disorder. In addition, we analyzed a subset of our pedigrees with psychotic features at this locus. We failed to find any association between the (CTG)n NOTCH4 polymorphism and either the bipolar or the psychotic bipolar phenotype in our 65 pedigrees.

The NOTCH4 locus is associated with susceptibility to schizophrenia.

Linkage disequilibrium mapping of the MHC region in 80 British parent-offspring trios showed that NOTCH4 was highly associated with schizophrenia. The A-->G substitution in the promoter region and the (CTG)n repeat in exon 1 of NOTCH4 may be candidate sites conferring susceptibility to schizophrenia.

Cloning, Characterization, and the Complete 56.8-Kilobase DNA Sequence of the Human NOTCH4 Gene

The first complete mammalian genomic sequence reported thus far in the Notch gene family, including a putative promoter region and 30 exons of the human NOTCH4 gene spanning 56.8 kb of DNA, were sequenced. The NOTCH4 locus contains a TATA-less promoter with two putative transcription initiation sites (Inr), three RBP-Jκ sites, and two GATA recognition sites. Two cDNA isoforms, NOTCH4(L) and NOTCH4(S),were identified. Whereas the NOTCH4(S) isoform contains the entire coding sequence, the NOTCH4(L) isoform has two unspliced intronic sequences between exons 11 and 12 and exons 20 and 21 and a misspliced exon 6. Consistent with these results, two alternatively spliced isoforms of transcripts of approximately 9.3 and 6.7 kb were detected by Northern blot analysis. The predicted amino acid sequence of the NOTCH4 protein based on the NOTCH4(S) cDNA sequence contains 2003 amino acids and includes the predominant motifs of the Notch family: 29 epidermal growth factor (EGF)-like repeats, 3 Notch/lin-12 repeats, a transmembrane region, 6 cdc10/Ankyrin repeats, and a PEST domain.