Health and population effects of rare gene knockouts in adult humans with related parents.

Examining complete gene knockouts within a viable organism can inform on gene function. We sequenced the exomes of 3222 British adults of Pakistani heritage with high parental relatedness, discovering 1111 rare-variant homozygous genotypes with predicted loss of function (knockouts) in 781 genes. We observed 13.7% fewer homozygous knockout genotypes than we expected, implying an average load of 1.6 recessive-lethal-equivalent loss-of-function (LOF) variants per adult. When genetic data were linked to the individuals' lifelong health records, we observed no significant relationship between gene knockouts and clinical consultation or prescription rate. In this data set, we identified a healthy PRDM9-knockout mother and performed phased genome sequencing on her, her child, and control individuals. Our results show that meiotic recombination sites are localized away from PRDM9-dependent hotspots. Thus, natural LOF variants inform on essential genetic loci and demonstrate PRDM9 redundancy in humans.

Genetic variants conferring susceptibility to Alzheimer's disease in the general population; do they also predispose to dementia in Down's syndrome.

BACKGROUND: Down's syndrome (DS) is caused by either complete or partial triplication of chromosome 21, affecting approximately 1/1000 live births, and it is widely accepted that individuals with DS are more likely to develop dementia of Alzheimer's disease (DAD) compared with the general population. Recent collaborative genome-wide association studies of large case control data sets of individuals with and without Alzhemier's disease (AD) have revealed new risk variants for dementia, as well as confirming previously identified risk variants. In this study, nine AD-derived SNPs, near or within the CR1 (rs3818361), BIN1 (rs744373), CD2AP (rs9349407), EPHA1 (rs11767557), CLU (rs1532278), MS4A6A/4A (rs610932), PICALM (rs561655), ABCA7 (rs3764650) and CD33 (rs3865444) genes were genotyped in 295 individuals with DS. RESULTS: There were no significant associations between these nine GWAS-derived SNPs and DAD in British Caucasian individuals with DS. Interestingly the CR1 rs3818361 variant appeared to be associated with mortality in our cohort, particularly in the subjects without dementia. To our knowledge, this is the first time that this variant has been implicated as a determinant of mortality and the finding warrants further investigation in other cohorts with DS. CONCLUSIONS: This study shows negative associations of nine AD-derived SNPs with DAD in DS. This may be due to the modest size of our cohort, which may indicate that our study is insufficiently powered to pick up such associations. We cannot conclusively exclude a role for these SNPs in DAD in DS. Clearly, efforts to investigate genetic variants with small effects on disease risk require a much larger cohort of individuals with DS. In fact, we hypothesize that a sample size of 4465 individuals with DS would be needed to determine the role in DAD in DS of the nine AD-derived SNPs investigated in this study. We therefore recommend that all national and international clinics with access to individuals with DS should contribute DNA samples to form DS consortia.

Genome-wide association study identifies a novel locus contributing to type 2 diabetes susceptibility in Sikhs of Punjabi origin from India.

We performed a genome-wide association study (GWAS) and a multistage meta-analysis of type 2 diabetes (T2D) in Punjabi Sikhs from India. Our discovery GWAS in 1,616 individuals (842 case subjects) was followed by in silico replication of the top 513 independent single nucleotide polymorphisms (SNPs) (P < 10⁻³) in Punjabi Sikhs (n = 2,819; 801 case subjects). We further replicated 66 SNPs (P < 10⁻4) through genotyping in a Punjabi Sikh sample (n = 2,894; 1,711 case subjects). On combined meta-analysis in Sikh populations (n = 7,329; 3,354 case subjects), we identified a novel locus in association with T2D at 13q12 represented by a directly genotyped intronic SNP (rs9552911, P = 1.82 × 10⁻8) in the SGCG gene. Next, we undertook in silico replication (stage 2b) of the top 513 signals (P < 10⁻³) in 29,157 non-Sikh South Asians (10,971 case subjects) and de novo genotyping of up to 31 top signals (P < 10⁻4) in 10,817 South Asians (5,157 case subjects) (stage 3b). In combined South Asian meta-analysis, we observed six suggestive associations (P < 10⁻5 to < 10⁻7), including SNPs at HMG1L1/CTCFL, PLXNA4, SCAP, and chr5p11. Further evaluation of 31 top SNPs in 33,707 East Asians (16,746 case subjects) (stage 3c) and 47,117 Europeans (8,130 case subjects) (stage 3d), and joint meta-analysis of 128,127 individuals (44,358 case subjects) from 27 multiethnic studies, did not reveal any additional loci nor was there any evidence of replication for the new variant. Our findings provide new evidence on the presence of a population-specific signal in relation to T2D, which may provide additional insights into T2D pathogenesis.

Circadian gene variants and susceptibility to type 2 diabetes: a pilot study.

BACKGROUND: Disruption of endogenous circadian rhythms has been shown to increase the risk of developing type 2 diabetes, suggesting that circadian genes might play a role in determining disease susceptibility. We present the results of a pilot study investigating the association between type 2 diabetes and selected single nucleotide polymorphisms (SNPs) in/near nine circadian genes. The variants were chosen based on their previously reported association with prostate cancer, a disease that has been suggested to have a genetic link with type 2 diabetes through a number of shared inherited risk determinants. METHODOLOGY/PRINCIPAL FINDINGS: The pilot study was performed using two genetically homogeneous Punjabi cohorts, one resident in the United Kingdom and one indigenous to Pakistan. Subjects with (N = 1732) and without (N = 1780) type 2 diabetes were genotyped for thirteen circadian variants using a competitive allele-specific polymerase chain reaction method. Associations between the SNPs and type 2 diabetes were investigated using logistic regression. The results were also combined with in silico data from other South Asian datasets (SAT2D consortium) and white European cohorts (DIAGRAM+) using meta-analysis. The rs7602358G allele near PER2 was negatively associated with type 2 diabetes in our Punjabi cohorts (combined odds ratio [OR] = 0.75 [0.66-0.86], p = 3.18 × 10(-5)), while the BMAL1 rs11022775T allele was associated with an increased risk of the disease (combined OR = 1.22 [1.07-1.39], p = 0.003). Neither of these associations was replicated in the SAT2D or DIAGRAM+ datasets, however. Meta-analysis of all the cohorts identified disease associations with two variants, rs2292912 in CRY2 and rs12315175 near CRY1, although statistical significance was nominal (combined OR = 1.05 [1.01-1.08], p = 0.008 and OR = 0.95 [0.91-0.99], p = 0.015 respectively). CONCLUSIONS/SIGNIFICANCE: None of the selected circadian gene variants was associated with type 2 diabetes with study-wide significance after meta-analysis. The nominal association observed with the CRY2 SNP, however, complements previous findings and confirms a role for this locus in disease susceptibility.

Effects of 16 genetic variants on fasting glucose and type 2 diabetes in South Asians: ADCY5 and GLIS3 variants may predispose to type 2 diabetes.

BACKGROUND: The Meta-Analysis of Glucose and Insulin related traits Consortium (MAGIC) recently identified 16 loci robustly associated with fasting glucose, some of which were also associated with type 2 diabetes. The purpose of our study was to explore the role of these variants in South Asian populations of Punjabi ancestry, originating predominantly from the District of Mirpur, Pakistan. METHODOLOGY/PRINCIPAL FINDINGS: Sixteen single nucleotide polymorphisms (SNPs) were genotyped in 1678 subjects with type 2 diabetes and 1584 normoglycaemic controls from two Punjabi populations; one resident in the UK and one indigenous to the District of Mirpur. In the normoglycaemic controls investigated for fasting glucose associations, 12 of 16 SNPs displayed ß values with the same direction of effect as that seen in European studies, although only the SLC30A8 rs11558471 SNP was nominally associated with fasting glucose (ß = 0.063 [95% CI: 0.013, 0.113] p = 0.015). Of interest, the MTNR1B rs10830963 SNP displayed a negative ß value for fasting glucose in our study; this effect size was significantly lower than that seen in Europeans (p = 1.29×10(-4)). In addition to previously reported type 2 diabetes risk variants in TCF7L2 and SLC30A8, SNPs in ADCY5 (rs11708067) and GLIS3 (rs7034200) displayed evidence for association with type 2 diabetes, with odds ratios of 1.23 (95% CI: 1.09, 1.39; p = 9.1×10(-4)) and 1.16 (95% CI: 1.05, 1.29; p = 3.49×10(-3)) respectively. CONCLUSIONS/SIGNIFICANCE: Although only the SLC30A8 rs11558471 SNP was nominally associated with fasting glucose in our study, the finding that 12 out of 16 SNPs displayed a direction of effect consistent with European studies suggests that a number of these variants may contribute to fasting glucose variation in individuals of South Asian ancestry. We also provide evidence for the first time in South Asians that alleles of SNPs in GLIS3 and ADCY5 may confer risk of type 2 diabetes.

Association of variants within APOE, SORL1, RUNX1, BACE1 and ALDH18A1 with dementia in Alzheimer's disease in subjects with Down syndrome.

BACKGROUND: Down syndrome (DS) is caused by either complete or partial triplication of chromosome 21, affecting approximately 1/1000 live births, and it is widely accepted that individuals with DS are more likely to develop dementia of Alzheimer's disease (DAD) compared with the general population. Many studies have investigated genetic susceptibility to AD in the general population, resulting in a number of potential candidate genes that may influence the development of DAD. The majority of these variants, however, have not been investigated in subjects with DS. AIM: The aim of this study was to determine whether genetic variants previously associated with AD in the general population, were also associated with DAD in individuals with DS. METHODS: Genotyping of 43 SNPs within 28 genes was undertaken in 187 individuals with Down syndrome with and without dementia of Alzheimer's disease, using the SNPlex platform. RESULTS: Significant associations of SNPs in five genes with DAD in DS were found, namely APOE, SORL1, BACE1, RUNX1 and ALDH18A1. As expected, the most strongly associated SNP was the APOE ɛ4 rs429358 variant (HR=2.47 [1.58, 3.87], p=7.52×10(-5)), although variants within the more recently implicated SORL1 and RUNX1 genes were also strongly associated with DAD in DS (HR=0.54 [0.37, 0.80], p=0.002 and HR=1.61 [1.15, 2.26], p=0.006 respectively). CONCLUSIONS: Our study demonstrates that a number of variants previously associated with AD in the general population are also associated with DAD in DS. To enable us to determine whether these variants, as well as other more recently revealed AD susceptibility variants, truly contribute to the development of DAD in DS, further multi-centre collaborative studies comprising large number of individuals with DS are needed.

The promoter polymorphism -232C/G of the PCK1 gene is associated with type 2 diabetes in a UK-resident South Asian population.

BACKGROUND: The PCK1 gene, encoding cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C), has previously been implicated as a candidate gene for type 2 diabetes (T2D) susceptibility. Rodent models demonstrate that over-expression of Pck1 can result in T2D development and a single nucleotide polymorphism (SNP) in the promoter region of human PCK1 (-232C/G) has exhibited significant association with the disease in several cohorts. Within the UK-resident South Asian population, T2D is 4 to 6 times more common than in indigenous white Caucasians. Despite this, few studies have reported on the genetic susceptibility to T2D in this ethnic group and none of these has investigated the possible effect of PCK1 variants. We therefore aimed to investigate the association between common variants of the PCK1 gene and T2D in a UK-resident South Asian population of Punjabi ancestry, originating predominantly from the Mirpur area of Azad Kashmir, Pakistan. METHODS: We used TaqMan assays to genotype five tagSNPs covering the PCK1 gene, including the -232C/G variant, in 903 subjects with T2D and 471 normoglycaemic controls. RESULTS: Of the variants studied, only the minor allele (G) of the -232C/G SNP demonstrated a significant association with T2D, displaying an OR of 1.21 (95% CI: 1.03 - 1.42, p = 0.019). CONCLUSION: This study is the first to investigate the association between variants of the PCK1 gene and T2D in South Asians. Our results suggest that the -232C/G promoter polymorphism confers susceptibility to T2D in this ethnic group. TRIAL REGISTRATION: UKADS Trial Registration: ISRCTN38297969.

Common variants of the TCF7L2 gene are associated with increased risk of type 2 diabetes mellitus in a UK-resident South Asian population.

BACKGROUND: Recent studies have implicated variants of the transcription factor 7-like 2 (TCF7L2) gene in genetic susceptibility to type 2 diabetes mellitus in several different populations. The aim of this study was to determine whether variants of this gene are also risk factors for type 2 diabetes development in a UK-resident South Asian cohort of Punjabi ancestry. METHODS: We genotyped four single nucleotide polymorphisms (SNPs) of TCF7L2 (rs7901695, rs7903146, rs11196205 and rs12255372) in 831 subjects with diabetes and 437 control subjects. RESULTS: The minor allele of each variant was significantly associated with type 2 diabetes; the greatest risk of developing the disease was conferred by rs7903146, with an allelic odds ratio (OR) of 1.31 (95% CI: 1.11 - 1.56, p = 1.96 x 10(-3)). For each variant, disease risk associated with homozygosity for the minor allele was greater than that for heterozygotes, with the exception of rs12255372. To determine the effect on the observed associations of including young control subjects in our data set, we reanalysed the data using subsets of the control group defined by different minimum age thresholds. Increasing the minimum age of our control subjects resulted in a corresponding increase in OR for all variants of the gene (p < or= 1.04 x 10(-7)). CONCLUSION: Our results support recent findings that TCF7L2 is an important genetic risk factor for the development of type 2 diabetes in multiple ethnic groups.

The MHC type 1 diabetes susceptibility gene is centromeric to HLA-DQB1.

HLA-DQB1 is widely considered to be the major histocompatibility complex (MHC) susceptibility gene for type 1 diabetes (T1D). However, since inheritance of the gene in T1D is recessive, the presence of the protective HLA-DQB1 0602 allele with normal nucleotide sequence in some patients raises the question of whether HLA-DQB1 is not the susceptibility locus itself but merely a good marker. HLA-DQB1 0602 is part of a conserved extended haplotype (CEH) [HLA-B7, SC31, DR2] (B7, DR2) with fixed DNA over more than 1Mb of genomic DNA that normally carries a protective allele at the true susceptibility locus. We postulated that, in patients with HLA-DQB1 0602, the protective allele at the susceptibility locus has been replaced by a susceptibility allele through an ancient crossover at meiosis centromeric to HLA-DQB1. We analyzed single nucleotide polymorphisms (SNPs) distinguishing the HLA-DQA2 (the first expressed gene centromeric to HLA-DQB1) allele on the normal HLA-B7, DR2 CEH from those on susceptibility CEHs in T1D patients and controls with HLA-DQB1 0602. All but 1 of 20 healthy control HLA-DQB1 0602 haplotypes had identical (consensus) first intron HLA-DQA2 5-SNP haplotypes. Fifteen of 19 patients with HLA-DQB1 0602 were homozygous for 1 or more HLA-DQA2 SNPs differing from consensus HLA-DQA2 SNPs, providing evidence of crossover involving the HLA-DQA2 locus. The remaining 4 patients were heterozygous at all positions and therefore uninformative. The loss of dominant protection usually associated with HLA-DQB1 0602 haplotypes is consistent with a locus centromeric to HLA-DQB1 being a major determinant of MHC-associated susceptibility, and perhaps the true T1D susceptibility locus.

A whole-genome admixture scan finds a candidate locus for multiple sclerosis susceptibility.

Multiple sclerosis is a common disease with proven heritability, but, despite large-scale attempts, no underlying risk genes have been identified. Traditional linkage scans have so far identified only one risk haplotype for multiple sclerosis (at HLA on chromosome 6), which explains only a fraction of the increased risk to siblings. Association scans such as admixture mapping have much more power, in principle, to find the weak factors that must explain most of the disease risk. We describe here the first high-powered admixture scan, focusing on 605 African American cases and 1,043 African American controls, and report a locus on chromosome 1 that is significantly associated with multiple sclerosis.

Analysis of the role of DPB1-encoded amino acids in the genetic predisposition to type I diabetes mellitus.

The role of the DPB1 gene in genetic susceptibility to type I diabetes has yet to be elucidated. Studies of DPB1 alleles are conflicting. Analysis at the amino acid level, rather than consideration of allelic polymorphism, has been informative in determining disease susceptibility encoded by the DRB1 and DQ genes. In this study, therefore, amino acid variation at polymorphic sites of the DPbeta peptide chain encoded by the second exon of the DPB1 gene was analyzed in diabetic and control subjects from white Caucasian, North Indian Asian, and Jamaican populations. Human leukocyte antigen genotypes and haplotypes were analyzed using a logistic-regression approach and the data were conditioned for the effects on disease risk of the DRB1, DQA1, and DQB1 genes. Eight DPbeta amino acid residues were significantly associated with type I diabetes independent of DR and DQ (DPbeta 9, 33, 35, 36, 55, 56, 57, and 69). None of these residues, however, correlated consistently with disease risk in all three racial groups. This contrasts with findings for the DRbeta, DQalpha and DQbeta peptide chains, where the identity of the amino acid at particular sites has been found to correlate with predisposition to type I diabetes.