Genome-wide linkage analysis and evidence of gene-by-gene interactions in a sample of 362 multiplex Parkinson disease families

Parkinson disease (PD) is the second most common neurodegenerative disorder. We studied 754 affected individuals, comprising 425 sibling pairs, to identify PD susceptibility genes. Screening of the parkin gene was performed in a subset of the sample having earlier age of PD onset or a positive LOD score with a marker in the parkin gene. All subjects were evaluated using a rigorous neurological assessment. Two diagnostic models were considered for genome-wide, non-parametric linkage analyses. Model I included only those individuals with a more stringent diagnosis of verified PD (216 sibling pairs) and resulted in a maximum LOD score of 3.4 on chromosome 2. Model II included all affected individuals (425 sibling pairs) and yielded a LOD score of 3.1 on the X chromosome. Our large sample was then employed to test for gene-by-gene (epistatic) interactions. A genome screen using the 23 families with PD patients having a mutation in only one allele of the parkin gene detected evidence of linkage to chromosome 10 (LOD=2.3). The 85 families with a very strong family history of PD were employed in a genome screen and, in addition to strong evidence of linkage to chromosome 2 (LOD=4.9), also produced a LOD of 2.4 on chromosome 14. A genome screen performed in the 277 families without a strong family history of PD detected linkage to chromosomes 10 (LOD=2.4) and X (LOD=3.2). These findings demonstrate consistent evidence of linkage to chromosomes 2 and X and also support the hypothesis that gene-by-gene interactions are important in PD susceptibility.     

Evidence for linkage disequilibrium between the alpha 7-nicotinic receptor gene (CHRNA7) locus and schizophrenia in Azorean families.

Recent studies have suggested that the alpha 7-nicotinic receptor gene (CHRNA7) may play a role in the pathogenesis of schizophrenia. The alpha 7-nicotinic receptor gene (CHRNA7) is involved in P50 auditory sensory gating deficits, and the genomic locus for this gene lies in the chromosome 15q13-14 regions. The human gene is partially duplicated (exons 5-10) with four novel upstream exons. The marker D15S1360 has been shown to be significantly linked with the phenotype of abnormal P50 suppression in schizophrenia families. The marker L76630 is 3 kb in the 3' direction from the last exon of the CHRNA7 gene and is located in the duplicated region. The function of the two L76630 copies is unknown. We genotyped three polymorphic markers D15S1360, D15S165, and L76630 that are localized in a genomic fragment containing the CHRNA7 in 31 Azorean schizophrenia families/trios (including 41 schizophrenia individuals and 97 unaffected families members). An overall analysis utilizing the family-based association test revealed significant linkage disequilibrium between L76630 and schizophrenia (P = 0.0004). Using the extended transmission disequilibrium test and limiting the analysis to one triad per family, transmission disequilibrium of D15S1360 was near significance (P = 0.078). The 15q13 region overlaps with the location of two well-known genomically imprinted disorders: Angelman syndrome and Prader-Willi syndrome. Therefore, we investigated maternal and paternal meioses. We found significant transmission disequilibrium for D15S1360 through paternal transmission (P = 0.0006) in our schizophrenia families. The L76630 marker showed a significant disequilibrium in maternal transmissions (P = 0.028). No parent-of-origin effect was found in D15S165. Overall, our results suggest that the CHRNA7 may play a role in schizophrenia in these families. A parent of origin effect may be present and requires further study.     

No association between 12 dopaminergic genes and schizophrenia in a large Dutch sample.

It has been suggested that genes involved in dopamine neurotransmission contribute to the pathogenesis of schizophrenia. However, reported associations of the disorder with genetic markers in dopaminergic genes have yielded inconsistent results. Possible explanations are differences in phenotyping, genetic heterogeneity, low marker informativity, and the use of small sample sizes. Here, we present a two-stage analysis of 12 dopaminergic genes in a large sample of Dutch schizophrenic patients. To reduce genetic heterogeneity, only patients with at least three Caucasian grandparents of Dutch ancestry were ascertained. An efficient genotyping strategy was used, in which polymorphic microsatellite markers were first screened for association in DNA pools. Promising results were followed up by individual genotyping in an extended sample. The pooled samples consisted of 208 schizophrenic patients and 288 unmatched control individuals. For each of the genes, more than one microsatellite marker was selected where possible, either intragenic or close to the gene. After correcting for multiple testing, significantly different allele frequencies were detected for DRD5 marker D4S615. Subsequently, we individually genotyped this particular marker and another DRD5 marker, as well as a DRD3 marker that could not be analyzed using the pooling strategy. This was done in an extended sample of 282 schizophrenic patients and a control sample of 585 individuals. In this second stage of the study, we found no association between these three markers and schizophrenia. The results of our comprehensive analysis provide no evidence for association between schizophrenia and 12 dopaminergic genes in a large Dutch sample.     

Examination of genetic linkage of chromosome 15 to schizophrenia in a large Veterans Affairs Cooperative Study sample.

Previous studies have reported genetic linkage evidence for a schizophrenia gene on chromosome 15q. Here, chromosome 15 was examined by genetic linkage analysis using 166 schizophrenia families, each with two or more affected subjects. The families, assembled from multiple centers by the Department of Veterans Affairs Cooperative Study Program, consisted of 392 sampled affected subjects and 216 affected sibling pairs. By DSM-III-R criteria, 360 subjects (91.8%) had a diagnosis of schizophrenia and 32 (8.2%) were classified as schizo-affective disorder, depressed. Participating families had diverse ethnic backgrounds. The largest single group were northern European American families (n = 62, 37%), but a substantial proportion was African American kindreds (n = 60, 36%). The chromosome 15 markers tested were spaced at intervals of approximately 10 cM over the entire chromosome and 2-5 cM for the region surrounding the alpha-7 nicotinic cholinergic receptor subunit gene (CHRNA7). These markers were genotyped and the data analyzed using semiparametric affecteds-only linkage analysis. In the European American families, there was a maximum Z-score of 1.65 between markers D15S165 and D15S1010. These markers are within 1 cM from CHRNA-7, the site previously implicated in schizophrenia. However, there was no evidence for linkage to this region in the African America kindreds.     

Evidence for linkage between the loci for transferrin and ceruloplasmin in man

Lod score analysis indicates probable linkage between the loci for transferrin and ceruloplasmin at a recombination frequency of approximately 10–15%. This human linkage relationship is homologous to one previously reported for cattle.     

Evidence for association between novel polymorphisms in the PRODH gene and schizophrenia in a Chinese population.

Haploinsufficiency for or mutation in at least one gene from the velocardiofacial syndrome (VCFS) region at chromosome 22q11 is implicated in psychosis. Linkage disequilibrium mapping of the region in patients identified a segment containing two genes, proline dehydrogenase (PRODH) and DGCR6, as candidates [Liu et al., 2002a] and by analysis of additional polymorphisms the PRODH gene was associated with schizophrenia in adult and early onset patients. In the present study we provide additional evidence in support of genetic association between PRODH and schizophrenia in a Chinese population. We analyzed the PRODH gene in a samples of schizophrenic patients and their families from Sichuan, SW China consisting of 528 family trios and sibling pairs. We genotyped six SNPs, PRODH*1195C-->T, PRODH*1482C-->T, PRODH*1483A-->G, PRODH*1766A-->G, PRODH*1852G-->A PRODH*1945T-->C, two of which (PRODH*1483A-->G and PRODH*1852G-->A) have not been previously reported. We found association with schizophrenia for two haplotypes consisting of PRODH*1945T-->C and PRODH*1852G-->A (Global P = 0.006), and PRODH*1852G-->A and PRODH*1766A-->G (Global P = 0.01) which include one of the newly identified markers. After six-fold Bonferroni correction for multiple testing the PRODH*1945T-C/PRODH*1852G-A haplotypes remained significant. This is a sub-haplotype of the PRODH haplotype previously associated with schizophrenia and it also maps to the 3' region of the gene, indicating that this is the region most likely to contain the underlying risk alleles. Overall this finding supports a role for the PRODH locus in schizophrenia.     

Evidence for transmission disequilibrium at the DAOA gene locus in a schizophrenia family sample

Recently, the DAOA gene locus on chromosome 13q32–q34 has been implicated in the etiology of schizophrenia. We genotyped three single-nucleotide polymorphisms (SNPs: rs778294, rs779293 and rs3918342) in this region in 126 Chinese family trios. In this study, we have identified statistically significant transmission disequilibrium in two markers rs778293 (P = 0.01) and rs3918342 (P = 0.02), and a highly significant under-transmission between haplotype CAT (P = 0.0005) and schizophrenia. The results provide further evidence to support that DAOA gene locus is involved in conferring susceptibility to schizophrenia.     

Autosomal linkage analysis of a Japanese single multiplex schizophrenia pedigree reveals two candidate loci on chromosomes 4q and 3q.

We analyzed a large multiplex schizophrenia pedigree collected in mid-eastern Japan using 322 microsatellite markers distributed throughout the whole autosome. Under an autosomal-dominant inheritance model, the highest pairwise LOD score (LOD = 1.69) was found at 4q (D4S2431: theta = 0.0), and LOD scores at two other loci 3q (ATA34G06) and 8q (D8S1128) were 1.62 and 1.46, respectively. In multipoint analysis, LOD scores of the regions on 4q and 3q remained at a similar level; however, the LOD score of the region on 8q apparently decreased. Additional dense map analysis revealed haplotypes on 4q and 3q regions shared by affected individuals. On chromosome 4q, the haplotype spanning about 8 centiMorgans (cM) was shared by four of six genotyped individuals with schizophrenia and one affected individual whose haplotype was estimated. On 3q, the haplotype spanning about 20 cM was shared by five genotyped individuals with schizophrenia. We obtained two candidate regions of major susceptibility loci for schizophrenia on chromosomes 3q and 4q.     

T-cell activation. I. Evidence for a functional linkage between class I MHC antigens and the Tc-Ti complex

This paper examines the possibility of a functional linkage between class I MHC molecules and the T-cell receptor complex for antigen (T3-Ti). A newly developed anti-CD3 antibody (500A2) was used as an activation signal for EL4 lymphoma cells and allospecific cytotoxic T-cell clones (CTL), and the production of IL-2/IL-2 receptor in EL4 cells and serine esterase in CTL was determined. Anti-CD3 antibody-induced activation of both EL4 and CTL cells was enhanced in the presence of immunologically cross-linked and immobilized anti-H-2 (class I) antibody reactive against the H-2 haplotype of the responding T cells. A number of H-2-negative and H-2-positive EL4 subclones were generated and tested for anti-CD3 antibody-induced IL-2/IL-2 receptor production. Although both H-2-positive and -negative subclones expressed CD3 antigen and produced IL-2 after activation with the phorbol ester TPA, only the H-2-positive cell clones produced IL-2 and expressed IL-2 receptor after anti-CD3 antibody induction. Our results are compatible with the existence of a functional linkage between the class I and the CD3 molecules on the surface of T cells.     

Report from the Maryland epidemiology schizophrenia linkage study: No evidence for linkage between schizophrenia and a number of candidate and other genomic regions using a complex dominant model

Our collaborative group has undertaken a linkage study of schizophrenia, using a systematic sample of patients admitted to Maryland hospitals. An initial sample of 39 families, each having two or more affecteds, was available for genotyping candidate genes, candidate regions, and highly polymorphic markers randomly distributed throughout the genome. We used a single complex dominant model (with a disease gene frequency of 0.005 and age-dependent penetrance for affected phenotype: for under 35, penetrance = .45; for 35 and older, penetrance = .85). We report here 130 markers, which met the exclusion criteria of LOD score <?2.00 at theta >0.01 in at least 10 informative families, and no evidence for heterogeneity. We also report here markers that were tested as candidates for linkage to the schizophrenic phenotype. They were selected based on the following criteria: (a) proximity to reported chromosomal rearrangements (both 5q and 11q), (b) suggestions of linkage from other families (5q), or (c) presence of a candidate gene (5q, 11q, 3q: Dopamine receptors 1, 2, and 3, respectively). We also tested for mutations of codon 717 in exon 17 of the amyloid precursor protein (APP) gene and were unable to detect the C to T substitution in our schizophrenic group. © 1994 Wiley-Liss, Inc.     

Evidence for linkage disequilibrium between the dopamine transporter and bipolar disorder

The mechanisms of hereditary deficiency of R binder, which originates in neutrophils and exocrine gland epithelium, are unknown and may be multiple. This led us to examine if defective R binder synthesis also involves proteins that colocalize with it in neutrophil-specific granules and exocrine epithelial cells and may be under common regulatory control. Stored plasma and saliva samples from five unrelated R binder-deficient patients and control subjects were assayed for R binder, lactoferrin, cationic antimicrobial protein-18, neutrophil gelatinase-associated lipocalin, gelatinase, lysozyme, and myeloperoxidase. One patient, patient A, had lactoferrin levels below the limits of detection in both plasma and saliva in addition to his R binder deficiency. Although his deficiency involved lactoferrin as well, he had no history of predisposition to infection. PCR amplification of his R binder gene promoter region and the beginning of the first exon revealed no DNA abnormalities. His son and the son of his equally deficient brother, both presumptive heterozygotes, had mild deficiency of both R binder and lactoferrin. The results show that R binder deficiency exists in at least two forms. One, presumably the less common of the two forms, is the new hereditary entity described here, which is characterized by deficiency of more than one specific granule protein in both plasma and saliva. Despite this more widely distributed absence of the proteins than is found in congenital specific granule deficiency, infection posed no clinical problem in the affected patient.     

An association study between polymorphism of L1CAM gene and schizophrenia in a Japanese sample.

L1CAM, a neural cell adhesion molecule, plays an important role in the development of the central nervous system. The human L1CAM gene is located in Xq28. Mutations in the gene are responsible for a wide spectrum of neurological abnormalities and mental retardation. Schizophrenia may result from early neurodevelopmental abnormalities. We screened 30 male and 30 female schizophrenic patients for their genomic sequence of the L1CAM gene in order to determine the DNA sequence variations. Three novel variations located in exon 18 (10564 G > A, GG/AA at codon 758), intron 11 (8575 A > C), and intron 25 (13504 C > T) were detected. An association study of the identified polymorphisms was then performed in a Japanese sample of 152 male and 115 female patients with schizophrenia and 121 male and 114 female control subjects. A statistically significant increase in the count of the 13504 T-allele was observed in the male patients, compared to the male controls, with no differences in the variations of exon 18 or intron 11. There was no statistically significant change in the distribution of allele or genotype of any variations in the female schizophrenics, in comparison with the female controls. These results suggest that the polymorphism in intron 25 plays a role in the genetic predisposition of male schizophrenia in the Japanese sample.     

Examination of genetic linkage of chromosome 15 to schizophrenia in a large Veterans Affairs Cooperative Study sample*

Previous studies have reported genetic linkage evidence for a schizophrenia gene on chromosome 15q. Here, chromosome 15 was examined by genetic linkage analysis using 166 schizophrenia families, each with two or more affected subjects. The families, assembled from multiple centers by the Department of Veterans Affairs Cooperative Study Program, consisted of 392 sampled affected subjects and 216 affected sibling pairs. By DSM‐III‐R criteria, 360 subjects (91.8%) had a diagnosis of schizophrenia and 32 (8.2%) were classified as schizo‐affective disorder, depressed. Participating families had diverse ethnic backgrounds. The largest single group were northern European American families (n = 62, 37%), but a substantial proportion was African American kindreds (n = 60, 36%). The chromosome 15 markers tested were spaced at intervals of approximately 10 cM over the entire chromosome and 2–5 cM for the region surrounding the α‐7 nicotinic cholinergic receptor subunit gene (CHRNA7). These markers were genotyped and the data analyzed using semiparametric affecteds‐only linkage analysis. In the European American families, there was a maximum Z‐score of 1.65 between markers D15S165 and D15S1010. These markers are within 1 cM from CHRNA‐7, the site previously implicated in schizophrenia. However, there was no evidence for linkage to this region in the African America kindreds. © 2001 Wiley‐Liss, Inc.     

Evidence for association and epistasis at the DAOA/G30 and D-amino acid oxidase loci in an Irish schizophrenia sample.

The D-amino acid oxidase (DAO) signaling pathway has been implicated in schizophrenia pathogenesis. This may be mediated through modulation of NMDA function by DAO, which is in turn activated by DAO activator (DAOA, formerly G72). Chumakov et al. (2002); PNAS 99: 13675-13680, identifying the novel schizophrenia susceptibility gene DAOA/G30 and a number of independent studies have since reported evidence of association between the DAOA and DAO genes and schizophrenia. However, at least two studies have failed to replicate the epistatic interaction between these loci described in the original report and there have been differences in the associated alleles/haplotypes reported at each locus. In this study, we performed association and epistasis analyses of the DAOA/G30 and DAO loci in a sample of 373 cases with DSM-IV schizophrenia/schizoaffective disorder and 812 controls from the Republic of Ireland. Corrected for the number of tests performed, we found evidence for association between markers at both genes and schizophrenia: DAOA/G30 (P = 0.005, OR = 1.34 (1.09, 1.65)) and DAO (P = 0.003, OR = 1.43 (1.12, 1.84). The data suggest that evidence for association at DAO (marker rs2111902) is more consistent than previously realized, particularly in Caucasian schizophrenia populations. We identified evidence for epistatic interaction between the associated SNPs at DAOA and DAO genes in contributing to schizophrenia risk (OR = 9.3 (1.4, 60.5). Based on these data, more systematic investigation of genes involved in DAO signaling is required.     

A study of linkage disequilibrium between polymorphic loci for monamine oxidases A and B in schizophrenia

Two X-linked microsatellites, (AC) n repeats at the monoamine oxidase (MAO)-A locus and (TG)n repeats at the MAO-B locus, were studied in 140 unrelated Caucasian male patients with schizophrenia and 91 unrelated Caucasian male controls. Among these subjects, we totally typed out nine alleles for the (AC) n repeats and eight alleles for the (TG) n repeats by using a PCR-based procedure. Allelic frequencies of either (AC) n repeats or (TG) n repeats were not found to be significantly different between patients and controls. However, a significant excess of the (AC)18/(TG)23 haplotype with a relative risk of 4.05 (95%; CI 1.15-14.26) was observed in patients with schizophrenia (Fisher's P = 0.011). The coefficient of linkage disequilibrium (delta) for the (AC)18/(TG)23 haplotype was 0.019 in schizophrenic patients and -0.046 in control subjects, respectively. The latter reached statistical significance (chi 2 = 6.02; df = 1; P < 0.02). The present findings suggest that linkage disequilibrium between polymorphic loci for human MAO-A and MAO-B may be associated with schizophrenia, and the (AC)18/(TG)23 haplotype may render an individual more vulnerable to such an illness.     

Evidence for the multigenic inheritance of schizophrenia.

Schizophrenia is assumed to have complex inheritance because of its high prevalence and sporadic familial transmission. Findings of linkage on different chromosomes in various studies corroborate this assumption. It is not known whether these findings represent heterogeneous inheritance, in which various ethnic groups inherit illness through different major gene effects, or multigenic inheritance, in which affected individuals inherit several common genetic abnormalities. This study therefore examined inheritance of schizophrenia at different genetic loci in a nationally collected European American and African American sample. Seventy-seven families were previously genotyped at 458 markers for the NIMH Schizophrenia Genetics Initiative. Initial genetic analysis tested a dominant model, with schizophrenia and schizoaffective disorder, depressed type, as the affected phenotype. The families showed one genome-wide significant linkage (Z = 3.97) at chromosome 15q14, which maps within 1 cM of a previous linkage at the alpha 7-nicotinic receptor gene. Chromosome 10p13 showed suggestive linkage (Z = 2.40). Six others (6q21, 9q32, 13q32, 15q24, 17p12, 20q13) were positive, with few differences between the two ethnic groups. The probability of each family transmitting schizophrenia through two genes is greater than expected from the combination of the independent segregation of each gene. Two trait-locus linkage analysis supports a model in which genetic alleles associated with schizophrenia are relatively common in the general population and affected individuals inherit risk for illness through at least two different loci.