Evaluation of IDDM8 susceptibility locus in a Russian simplex family data set

Type 1 diabetes (T1D) susceptibility locus, IDDM8, has been accurately mapped to 200 kilobases at the terminal end of chromosome 6q27. This is within the region which harbours a cluster of three genes encoding proteasome subunit beta 1 (PMSB1), TATA-box binding protein (TBP) and a homologue of mouse programming cell death activator 2 (PDCD2). In this study, we evaluated whether these genes contribute to T1D susceptibility using the transmission disequilibrium test of the data set from 114 affected Russian simplex families. The A allele of the G/A1180 single nucleotide polymorphism (SNP) at the PDCD2 gene, which was significant in its preferential transfer from parents to diabetic children (75 transmissions vs. 47 non-transmissions, chi2=12.85, P corrected=0.0038), was found to be associated with T1D. G/A1180 dimorphism and two other SNPs, C/T771 TBP and G/T(-271) PDCD2, were shown to share three common haplotypes, two of which (A-T-G and A-T-T) have been associated with higher development risk of T1D. The third haplotype (G-T-G) was related to having a lower risk of disease. These findings suggest that the PDCD2 gene is a likely susceptibility gene for T1D within IDDM8. However, it was not possible to exclude the TBP gene from being another putative susceptibility gene in this region.     

The TAF5L gene on chromosome 1q42 is associated with type 1 diabetes in Russian affected patients

Type 1 diabetes (T1D) is a multifactorial autoimmune disease, with strong genetic component. Several susceptibility loci contribute to genetic predisposition to T1D. One of these loci have been mapped to chromosome 1q42 in UK and US joined affected family data sets but needs to be replicated in other populations. In this study, we evaluated sixteen microsatellites located on 1q42 for linkage with T1D in 97 Russian affected sibling pairs. A 2.7-cm region of suggestive linkage to T1D between markers D1S1644 and D1S225 was found by multipoint linkage analysis. The peak of linkage was shown for D1S2847 (P = 0.0005). Transmission disequilibrium test showed significant undertransmission of the 156-bp allele of D1S2847 from parents to diabetic children (28 transmissions vs. 68 nontransmissions, P = 0.043) in Russian affected families. A preferential transmission from parents to diabetic offspring was also shown for the T(-25) and T1362 alleles of the C/T(-25) and C/T1362 dimorphisms, both located at the TAF5L gene, which is situated 103 kb from D1S2847. Together with the A/C744 TAF5L SNP, these markers share common T(-25)/A744/T1362 and C(-25)/C744/T1362 haplotypes associated with higher and lower risk of diabetes (Odds Ratio = 2.15 and 0.62, respectively). Our results suggest that the TAF5L gene, encoding TAF5L-like RNA polymerase II p300/CBP associated factor (PCAF)-associated factor, could represent the susceptibility gene for T1D on chromosome 1q42 in Russian affected patients.     

KCNQ4: a gene for age‐related hearing impairment?

Age-related hearing impairment (ARHI) is the most common sensory impairment among the elderly. It is a complex disorder influenced by genetic as well as environmental factors. SNPs in a candidate susceptibility gene, KCNQ4, were examined in two independent Caucasian populations. Two quantitative trait locus (QTL) values were investigated: Zhigh and Zlow, a measure of high and respectively low frequency hearing loss. In the first population, the statistical analysis of 23 genotyped SNPs spread across KCNQ4 resulted in significant p-values for two SNPs for Zhigh-SNP9 (NT_004511:g.11244177A > T) and SNP15 (NT_004511:g.11257005C > T; NP_004691:p.Ala259Ala), and one SNP for Zlow-SNP12 (NT_004511:g.11249550A > T). The linkage disequilibrium (LD) structure of KCNQ4 was subsequently determined in a 34-kb region surrounding the significant SNPs, resulting in three LD-blocks. LD-block 1 contains SNP9 and covers an area of 5 kb, LD-block 2 measures 5 kb and surrounds SNP13 (NT_004511:g.11253513A > G) to SNP18 (NT_004511:g.11257509G > A; NP_004691:p.Thr293Thr), and LD-block 3 spans 7 kb. Five tag-SNPs of block 1 and 2, and 2 extra SNPs were subsequently genotyped in the second population. Again, several SNPs were positively associated with ARHI: one SNP (SNP18) for the high frequencies and three SNPs (SNP9, SNP12, and SNP18) for the low frequencies, although only a single SNP (SNP12) resulted in significant p-values in both populations. Nevertheless, the associated SNPs of both populations were all located in the same 13-kb region in the middle of the KCNQ4 gene.     

CT60 single nucleotide polymorphism of the CTLA-4 gene is associated with susceptibility to Graves' disease in the Taiwanese population

Graves' disease (GD) is an autoimmune disease but the underlying etiology has not been completely elucidated. Genetic susceptibility has been believed to play a major role. Recent studies showed that the CT60 single nucleotide polymorphism (SNP), which is in the 3'-noncoding region of the CTLA-4 gene, is strongly associated with some immune-mediated diseases. The aim of this study was to test for association between GD susceptibility and polymorphisms of CTLA-4 (ie, the CT60 SNP and the exon 1 +49 SNP) in the Taiwanese population. Our results demonstrate significant differences in the frequencies of the genotypes and alleles between 107 GD patients and 101 control subjects in the CT60 and exon 1 +49 SNPs (p <0.05). Significant differences in phenotypes were only found for CT60 SNP (78.4% vs 67.8% between patients and controls; chi2 = 3.93, p = 0.047). Furthermore, we found that the G/G genotype of both CT60 and exon 1 +49 was associated with increased risk for GD (p = 0.022, OR = 1.97). Significant linkage disequilibrium was found between the CT60 SNP and the exon 1 +49 SNP in both GD patients and control subjects (D' = 1.00). Because of tight linkage disequilibrium, a combination of these SNPs enhanced the role of the CTLA-4 gene in GD. The frequency of the disease-susceptible G allele of CT60 was comparable to that in Japanese and higher than in Caucasians. In conclusion, we provide evidence that CT60 SNP is associated with susceptibility to GD in the Taiwanese population.     

Type 2 deiodinase and host responses of sepsis and acute lung injury

The role of thyroid hormone metabolism in clinical outcomes of the critically ill remains unclear. Using preclinical models of acute lung injury (ALI), we assessed the gene and protein expression of type 2 deiodinase (DIO2), a key driver for synthesis of biologically active triiodothyronine, and addressed potential association of DIO2 genetic variants with ALI in a multiethnic cohort. DIO2 gene and protein expression levels in murine lung were validated by microarrays and immunoblotting. Lung injury was assessed by levels of bronchoalveolar lavage protein and leukocytes. Single-nucleotide polymorphisms were genotyped and ALI susceptibility association assessed. Significant increases in both DIO2 gene and D2 protein expression were observed in lung tissues from murine ALI models (LPS- and ventilator-induced lung injury), with expression directly increasing with the extent of lung injury. Mice with reduced levels of DIO2 expression (by silencing RNA) demonstrated reduced thyroxine levels in plasma and increased lung injury (increased bronchoalveolar lavage protein and leukocytes), suggesting a protective role for DIO2 in ALI. The G (Ala) allele of the Thr92Ala coding single-nucleotide polymorphism (rs225014) was protective in severe sepsis and severe sepsis-associated ALI after adjustments for age, sex, and genetic ancestry in a logistic regression model in European Americans. Our studies indicate that DIO2 is a novel ALI candidate gene, the nonsynonymous Thr92Ala coding variant of which confers ALI protection. Increased DIO2 expression may dampen the ALI inflammatory response, thereby strengthening the premise that thyroid hormone metabolism is intimately linked to the integrated response to inflammatory injury in critically ill patients.     

Transmission disequilibrium and haplotype analyses of the G72/G30 locus: suggestive linkage to schizophrenia in Palestinian Arabs living in the North of Israel.

Association of the G72/G30 locus with schizophrenia was recently reported in French Canadian, Russian, and Ashkenazi populations using case-control studies. In the present study we hypothesize the existence of a G72/G30 risk allele over-transmitted to affected sibs in Palestinian Arab families. A total of 223 Palestinian Arab families that included an affected offspring and parents were genotyped with 11 SNPs encompassing the G72/G30 genes. The families were recruited from three regions of Israel: 56 from the North (Afula), 136 from the central hill region (Bethlehem, Palestinian Authority), and 31 from the South (Beersheva). Individual SNP analyses disclosed a risk allele in SNP rs3916970 by both haplotype relative risk (HRR: chi(2) = 5.59, P = 0.018) and transmission disequilibrium test (TDT: chi(2) = 6.03, P = 0.014) in the Afula families. Follow-up multilocus analysis using family-based association tests (FBAT: z = 2.197, P = 0.028) exposed the adjacent haplotype. SNP rs3916970 is located about 8 kb from the linkage disequilibrium block that was reported to be associated with schizophrenia in Ashkenazi Jews. Excess of similar haplotypes of this region was observed in the Palestinian Arabs and the Ashkenazi patients. These data suggest a common risk factor for schizophrenia susceptibility in the G72/G30 locus among Ashkenazi Jews and Palestinian Arabs. The results strengthen previous reports on the role of this locus in the etiology of schizophrenia.     

Identification of DIO2 as a new susceptibility locus for symptomatic osteoarthritis

Osteoarthritis [MIM 165720] is a common late-onset articular joint disease for which no pharmaceutical intervention is available to attenuate the cartilage degeneration. To identify a new osteoarthritis susceptibility locus, a genome-wide linkage scan and combined linkage association analysis were applied to 179 affected siblings and four trios with generalized osteoarthritis (The GARP study). We tested, for confirmation by association, 1478 subjects who required joint replacement and 734 controls in a UK population. Additional replication was tested in 1582 population-based females from the Rotterdam study that contained 94 cases with defined hip osteoarthritis and in 267 Japanese females with symptomatic hip osteoarthritis and 465 controls. Suggested evidence for linkage in the GARP study was observed on chromosome 14q32.11 (log of odds = 3.03, P = 1.9 x 10(-4)). Genotyping tagging single-nucleotide polymorphisms covering three important candidate genes revealed a common coding variant (rs225014; Thr92Ala) in the iodothyronine-deiodinase enzyme type 2 (D2) gene (DIO2 [MIM 601413]) which significantly explained the linkage signal (P = 0.006). Confirmation and replication by association in the additional osteoarthritis studies indicated a common DIO2 haplotype, exclusively containing the minor allele of rs225014 and common allele of rs12885300, with a combined recessive odds ratio of 1.79, 95% confidence interval (CI) 1.37-2.34 with P = 2.02 x 10(-5) in female cases with advanced/symptomatic hip osteoarthritis. The gene product of this DIO2 converts intracellular pro-hormone-3,3',5,5'-tetraiodothyronine (T4) into the active thyroid hormone 3,3',5-triiodothyronine (T3) thereby regulating intracellular levels of active T3 in target tissues such as the growth plate. Our results indicate a new susceptibility gene (DIO2) conferring risk to osteoarthritis.     

Genetic Variation at the Chromosome 16 Chemokine Gene Cluster: Development of a Strategy for Association Studies in Complex Disease

The chemokine gene cluster [CCL22, CX3CL1, CCL17] (previously known as [SCYA22, SCYD1, SCYA17]) is a candidate locus for one of the susceptibility genes for inflammatory bowel disease that are located in the peri‐centromeric region of chromosome 16. Screening for sequence variation at this locus led to the detection of 14 single nucleotide polymorphisms (SNPs). An efficient experimental and computational approach was developed to estimate allele frequencies and pairwise linkage disequilibrium relationships between SNPs at this locus, and to test them for association with inflammatory bowel disease. The 12 common SNPs were assigned to 5 distinct linkage disequilibrium groups. Genotyping of one SNP from each linkage disequilibrium group in a large cohort of families with inflammatory bowel disease did not provide convincing evidence of association with either Crohn's disease or ulcerative colitis. We describe an efficient experimental design from SNP screening to association testing. This strategy can be used to test candidate genes for involvement in susceptibility to complex disease.     

Identification of independent risk loci for Graves' disease within the MHC in the Japanese population

To identify genetic variants that confer the risk of Graves' disease (GD) in the Japanese population, we conducted a two-stage genome-wide association study (GWAS) using 1119 Japanese individuals with GD and 2718 unrelated controls, and a subsequent replication study using independent 432 GD cases and 1157 controls. We identified 34 single nucleotide polymorphisms (SNPs) to be significantly associated with GD in the GWAS phase. Twenty-two out of 34 SNPs remained positive in the replication study. All 22 SNPs were located within the major histocompatibility complex (MHC) locus on chromosome 6p21. No strong long-range linkage disequilibrium (LD) was observed among the 22 SNPs, indicating independent involvement of multiple loci within the MHC with the risk of GD. Multivariate stepwise logistic regression analysis selected rs3893464, rs4313034, rs3132613, rs4248154, rs2273017, rs9394159 and rs4713693, as markers for independent risk loci for GD. The analysis of LD between these seven SNPs and tagging SNPs for GD-associated human leukocyte antigen (HLA) alleles in the Japanese population (HLA-DPB1(*)0501 and HLA-A(*)0206) demonstrated that all of and five of seven SNPs were not in strong LD with HLA-DPB1(*)0501 and HLA-A(*)0206, respectively. Although causal variants remain to be identified, our results demonstrate the existence of multiple GD susceptibility loci within the MHC region.     

Association of NOD2 with Crohn's disease in a homogenous Irish population

Linkage of IBD to the pericentromeric region of chromosome 16 has been widely confirmed by analyses of multiple populations. The NOD2 gene is located in the peak region of linkage on chromosome 16 and thought to be involved in the activation of nuclear factor (NF) kappaB in response to bacterial components. Mutations in the NOD2 gene are found to be strongly associated with susceptibility to Crohn's disease (CD). A total of 65 Irish CD families were genotyped to determine if NOD2 mutations conferred susceptibility to CD and the prevalence of these mutations in sporadic and familial forms of the disease. The Irish population is relatively homogenous and thus may provide advantages in genetic studies of complex diseases. We confirmed the IBD1 locus as a susceptibility locus for IBD within the Irish population by linkage analysis followed by linkage disequilibrium studies. No significant evidence of linkage was observed to the previously identified regions on chromosomes 1, 12 and 14. In all, 131 CD affected families were then genotyped for seven of the previously published NOD2 single-nucleotide polymorphisms (SNPs). Allelic transmission distortion was investigated using the pedigree disequilibrium test (PDT). SNP13 (3020insC) was found to be associated with CD (P=0.0186). Patients who possessed a rare allele of SNP8, 12 or 13 presented earlier when compared to patients without rare variants (mean age, 20.1 vs 24 years, P=0.011) and the rare allele of SNP13 was observed to be predominantly linked to ileal disease (P=0.02). This report confirms the importance of NOD2 as a susceptibility gene for CD within the Irish population.     

Thyroglobulin polymorphisms in Tunisian patients with autoimmune thyroid diseases (AITD)

The thyroglobulin (Tg) gene was reported to be linked and/or associated to autoimmune thyroid diseases (AITD) development in European Caucasian populations. Here, we attempt to replicate this finding and to evaluate the contribution of the Tg gene in the genetic susceptibility of AITD in the Tunisian population. We examined the genomic region (11.5cM) containing the Tg gene by genotyping seven microsatellite markers and four SNPs located respectively at exon 10 (Ser715Ala), exon 12 (Met1009Val), exon 21 (Ala1483Ala) and exon 33 (Arg1980Trp) in 15 Tunisian multiplex families affected with AITD including Graves' disease (GD) and Hashimoto's thyroiditis (HT) (members: 87; patients: 27 GD and 16 HT). A case-control study was performed by genotyping the Tgms2 intragenic microsatellite marker (intron 27) and four intragenic SNPs on 108 unrelated patients affected with GD and 169 normal controls. Analysis of family data did not show linkage of the thyroglobulin gene with AITD nor did analysis of case-control data show association of Tgms2 or SNPs with GD. In contrast to the European Caucasian population, we failed to detect any contribution of Tg gene in the genetic component of Tunisian AITD.     

Identification of susceptibility loci for autoimmune thyroid disease to 5q31-q33 and Hashimoto's thyroiditis to 8q23-q24 by multipoint affected sib-pair linkage analysis in Japanese.

Autoimmune thyroid disease (AITD), including Graves' disease (GD) and Hashimoto's thyroiditis (HT), is caused by multiple genetic and environmental factors. The clinical and immunological features of GD and HT are distinct; however, there are multiplex families with both GD and HT, and cases in which GD evolves into HT. Thus, there may be specific susceptibility loci for GD or HT, and common loci controlling the susceptibility to both GD and HT may exist. A genome-wide analysis of data on 123 Japanese sib-pairs affected with AITD was made in which GD- or HT-affected sib-pairs (ASPs) were studied to detect GD- or HT-specific susceptibility loci, and all AITD-ASPs were used to detect AITD-common susceptibility loci. Our study revealed 19 regions on 14 chromosomes (1, 2, 3, 5, 6, 8, 9, 10, 11, 12, 13, 15, 18 and 22) where the multipoint maximum LOD score (MLS) was >1. Especially, chromosome 5q31-q33 yielded suggestive evidence for linkage to AITD as a whole, with an MLS of 3.14 at D5S436, and chromosome 8q23-q24 yielded suggestive evidence for linkage to HT, with an MLS of 3.77 at D8S272. These observations suggest the presence of an AITD susceptibility locus at 5q31-q33 and a HT susceptibility locus at 8q23-q24.     

DIO2基因单核苷酸多态性与精神发育迟滞相关性研究

目的探讨DIO2基因多态性及单倍型与精神发育迟滞的相关关系。方法本研究以344个汉族精神发育迟滞患者及其亲属组成的家系为研究对象．在DIO2上选择了3个合适的SNPs作为标记．利用PCR—SSCP和PCR—RFLP方法完成基因分型。应用FBAT软件对多态位点及可能组成的单倍型与精神发育迟滞的关系进行分析。结果单个位点家系关联性分析结果提示：rs225015、rs225014和rs225012位点各等位基因从亲代传递给患病子代没有观察到显著性（P〉0．05）。多态性位点间的连锁不平衡检验显示：3个位点rs225015、rs225014和rs225012均具有较高的连锁不平衡（D’〉0．8）。单倍型分析结果显示：3个位点构成的单体型GTG在附加遗传模型（Z=2．226,P=0．026019）和隐性遗传模型（Z=2．651,P=0．008023）与MR相关。结论DIO2基因可能与精神发育迟滞的易感性有关。     

Family-based association study between autism and glutamate receptor 6 gene in Chinese Han trios.

The glutamate pathways are involved in diverse processes such as learning and memory, epilepsy, and they play important roles in neural plasticity, neural development, and neurodegeneration. It has been proposed that autism could be a hypoglutamatergic disorder. Recently, Jamain et al. reported that the glutamate receptor 6 (GluR6 or GRIK2) is in linkage disequilibrium with autism. In the present study, the transmission disequilibrium test (TDT) and the haplotype transmission were performed to analyze the four SNPs (SNP1: rs995640; SNP2: rs2227281; SNP3: rs2227283; SNP4: rs2235076) of GluR6 in 174 Chinese Han parent-offspring trios. The TDT demonstrated that the two SNPs (SNP2 and SNP3) showed preferential transmission (TDT P = 0.032). The global chi(2) test for haplotype transmission also revealed an association between GluR6 and autism (chi(2) = 10.78, df = 3, P = 0.013). Our results suggested that GluR6 is in linkage disequilibrium with autism.     

Molecular screening and association studies of retinoid-related orphan receptor gamma (RORC): a positional and functional candidate for type 2 diabetes

The retinoid-related orphan receptor gamma (RORC) is a member of the nuclear hormone superfamily which maps to the 1q21-q23 region. Linkage of type 2 diabetes (T2DM) to this region is well replicated. Several factors argue that RORC is a strong candidate for T2DM susceptibility within this region. RORC may form heterodimers with peroxisome-proliferator activated receptor gamma, it is expressed at high levels in skeletal muscle, and expression is induced in adipocytes during differentiation. To test the hypothesis that sequence variation in RORC is a risk factor for T2DM, we screened approximately 21kb of DNA for sequence variation, including 11 exons of the RORC gene, a region 1-kb upstream (5' flanking region), intronic regions flanking the exons, and the entire 3' untranslated region (UTR). Screening was performed using single strand conformation polymorphism (SSCP) analysis in Caucasian individuals of northern European ancestry and in African American individuals. We detected 11 single nucleotide polymorphisms (SNPs), ranging from the promoter region to intron 10. We also confirmed 2 SNPs from public databases that were in regions not included in our screening. Only 1 SNP was nonsynonymous, resulting in Ala to Gly at residue 464 (exon 10). All other SNPs were noncoding. One SNP (intron 3) was unique to Caucasians, and three SNPs (Ala464Gly, intron 2, intron 6) were specific to African American subjects. We typed 7 SNPs spanning the gene from the promoter to 3' UTR in unrelated cases with T2DM and controls of Northern European ancestry. We also tested linkage of a microsatellite within the RORC gene. Modest evidence for linkage (LOD=1.47) was seen on two-point analysis, but no linkage to the RORC region was found on multipoint analysis. However, transmission of the microsatellite alleles from parents to affected offspring showed a trend to deviate from the expected 50% (p=0.078). No association of any other SNP with T2DM was found, but the Ala454Gly variant was 3-fold more common among African American patients with diabetes than in controls. SNPs 1, 2 and 4 were in strong linkage disequilibrium (D>0.85) and may constitute a haplotype block. Our data suggest that RORC cannot explain the linkage of T2DM in this region. The role of the unusual Ala454Gly variant will require a much larger study size to evaluate.     

Evidence of association between FKBP1B and thyroid autoimmune disorders in a large Tunisian family

FKBP1B belongs to immunophilins superfamily and functions as a cytosolic receptor protein of FK506. The role of FKBP1B in the immunosuppressive pathway of FK506 is well established. Previously, we reported a strong evidence of linkage between D2S171 microsatellite marker (located in vicinity of FKBP1B gene) and susceptibility to autoimmune thyroid diseases (AITDs). In this study, we report linkage disequilibrium between the dimorphism (C/T) in the 3' untranslated region (3' UTR) of FKBP1B gene and susceptibility to AITDs. DNAs were extracted from a large Tunisian family affected with Graves' disease (GD) and Hashimoto's thyroiditis (HT) and analysed by PCR-RFLP using DraIII restriction enzyme. Our results showed an excess of transmission of the allele C from heterozygous parents to affected offspring (transmission disequilibrium test (TDT) = 4.76; p = 0.012). This suggests a linkage disequilibrium of 3' UTR (C/T) SNP with AITDs. Moreover, The FBAT analysis gives a significant association with the C allele under the recessive model (chi2 = 5.50; p = 0.018). These results support the involvement of FKBP1B gene in the genetic susceptibility to the AITDs development in the studied family.     

Genetic analysis and functional evaluation of the C/T(-318) and A/G(-1661) polymorphisms of the CTLA-4 gene in patients affected with Graves' disease

In this study, we evaluated the A/G(-1661), C/T(-318), A/G49 and A/G6230 single nucleotide polymorphisms (SNPs) of the cytotoxic T lymphocyte antigen 4 (CTLA-4) gene for association with Graves' disease (GD) in 126 Russian simplex families. The conditional TDT analysis revealed significant overtransmission of the A(-1661)G(-318) haplotype (P = 0.033) and undertransmission of the GT haplotype (P = 0.0043) from parents homozygous for both +49 and +6230 polymorphisms. Parents homozygous for both (-1661) and (-318) markers significantly overtransmitted the G49G6230 haplotype (P = 0.0013) and undertransmitted the AG haplotype (P = 0.035) to affected offspring. This suggests in favor of the independent genetic effects of the 3' and 5'ends of CTLA-4 in conferring the susceptibility to GD. Both SNPs located at the 5' untranslated region of CTLA-4 were functionally analyzed using the luciferase reporter assay. We observed differential activation of the C/T(-318) promoter variant when Jurkat T cells and HeLa cells were cotransfected with a plasmid expressing lymphoid enhancing factor 1 (LEF1) and various CTLA-4 promoter constructs. The (-318) SNP modifies a putative binding site for LEF1 so that it alters the stimulating effect of LEF1 on the expression ability of the CTLA-4 promoter. The (-1661) dimorphism modifies a potential binding site for myocyte enhancer factor 2 (MEF2). No significant correlation between the (-1661) SNP and MEF2 activity in cotransfection experiments was found. Observed data help for further understanding a functional role of CTLA-4 promoter polymorphisms in the pathogenic mechanism of GD.     

The cytotoxic T lymphocyte antigen-4 is a major Graves' disease locus.

Graves' disease (GD) is an autoimmune thyroid disorder that is inherited as a complex trait. We have genotyped 77 affected sib-pairs with autoimmune thyroid disease for eight polymorphic markers spanning the cytotoxic T lymphocyte antigen-4 ( CTLA-4 ) region of chromosome 2q31-q33, and for five markers spanning the major histocompatibility complex ( MHC ) region of chromosome 6p21. Non-parametric analysis showed linkage of GD to the CTLA-4 region with a peak non-parametric linkage (NPL) score of 3.43 ( P = 0.0004) at the marker D2S117. The proportion of affected full-sibs sharing zero alleles (z0) reached a minimum of 0.113 close to D2S117, giving a locus-specific lambdas for this region of 2.2. Families with brother-sister sib-pairs showed a peak NPL of 3.46 ( P = 0.0003, lambdas > 10) at D2S117, compared with 2.00 ( P = 0.02, lambdas = 1.9) in the families with only affected females, suggesting a stronger influence in families with affected males. Association between GD and the G allele of the Thr17Ala polymorphism within the CTLA-4 gene ( CTLA4A/G ) was observed using unaffected sib controls ( P = 0.005). Lesser evidence for linkage was found at the MHC locus, with a peak NPL score of 1.95 ( P = 0.026), between the markers D6S273 and TNFalpha. We demonstrate that the CTLA-4 locus (lambdas = 2.2) and the MHC locus (lambdas = 1.6) together confer approximately 50% of the inherited susceptibility to GD disease in our population.