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Our previous neurocognitive studies of schizophrenia outlined two clusters of affected subjects--cognitively spared (CS) and cognitive deficit (CD), the latter's characteristics pointing to developmental origins and impaired synaptic plasticity. Here we investigate the contribution of polymorphisms in major regulators of these processes to susceptibility to schizophrenia and to CD in patients. We examine variation in genes encoding proteins at the gateway of Reelin signaling: ligands RELN and APOE, their common receptors APOER2 and VLDLR, and adaptor DAB1. Association analysis with disease outcome and cognitive performance in the Western Australian Family Study of Schizophrenia (WAFSS) was followed by replication analysis in the Australian Schizophrenia Research Bank (ASRB) and in the Health in Men Study (HIMS) of normal aging males. In the WAFSS sample, we observed significant association of APOE, APOER2, VLDLR, and DAB1 SNPs with disease outcome in the case-control and CD-control datasets, and with pre-morbid intelligence and verbal memory in cases. HIMS replication analysis supported rs439401 (APOE regulatory region), and rs2297660 and rs3737983 (APOER2), with an effect on memory performance in normal aging subjects consistent with the findings in schizophrenia cases. APOER2 gene expression analysis revealed lower transcript levels in lymphoblastoid cells from cognitively impaired schizophrenia patients of the alternatively spliced exon 19, mediating Reelin signaling and synaptic plasticity in the adult brain. ASRB replication analysis produced marginally significant results, possibly reflecting a recruitment strategy biased toward CS patients. The data suggest a contribution of neurodevelopmental/synaptic plasticity genes to cognitive impairment in schizophrenia.

In a previous study we identified a relatively homogeneous subtype of schizophrenia characterized by pervasive cognitive deficit, which was the exclusive contributor to our findings of linkage to 6p25-p24. The 6p region contains Dysbindin (DTNBP1), considered to be one of the major schizophrenia candidate genes. While multiple studies have reported association between genetic variation in DTNBP1 and schizophrenia, the findings have been inconsistent and controversial, leading to recent calls for systematic re-examination and unambiguous evidence of association. To address this, we have undertaken a comprehensive survey of common genetic variation within DTNBP1 and its association with schizophrenia, using a HapMap-based gene-tagging approach. We genotyped 39 tSNPs in a sample of 336 cases and 172 controls of Anglo-Irish ancestry, with the phenotype defined as clinical schizophrenia, and as composite neurocognitive endophenotypes. Allele and haplotype frequencies, and LD structure in our control sample were similar to those in other European populations. Using multivariate generalized linear modeling, we observed no significant association between any tSNP and any outcome variable. Association with haplotypes was examined across the gene and in the previously associated 5' region. Neither global haplotype tests, nor specific analysis of the "risk" haplotype previously reported in an ethnically related population, the Irish high-density schizophrenia families, showed significant evidence of association with schizophrenia or with the neurocognitive endophenotypes in our sample. The framework and results of this study should facilitate further attempts at re-analysis of DTNBP1, in terms of standardized approaches to both phenotype definition and analysis of genetic variation. (C) 2008 Wiley-Liss, Inc.

PARL (presenilin-associated rhomboid-like) is a mitochondrial protein involved in mitochondrial membrane remodelling, and maps to a quantitative trait locus (3q27) associated with metabolic traits. Recently the rs3732581 (Leu262Val) variant was found to be associated with increased levels of plasma insulin, a finding not replicated in a larger cohort. The aim of the current study was to investigate the associations between rs3732581 and levels of plasma insulin, metabolic syndrome (MetS) and its components, and cardiovascular disease. The CUPID population consisted of 556 subjects with angiographically proven CAD and the CUDAS cohort consisted of 1,109 randomly selected individuals from Perth, Western Australia. Samples were genotyped using mutation-specific PCR. No significant associations were observed between rs3732581 and levels of plasma insulin, glucose, BMI or MetS in either population. However, carriers of the minor allele had significantly lower mean intima-media thickness (IMT) [0.69 mm, 95% CI (0.69, 0.70 mm); P = 0.004], compared with major allele homozygotes [mean IMT = 0.71 mm, 95% CI (0.70, 0.72 mm)] in the CUDAS population. Further analysis using a recessive model showed homozygous carriers of the minor allele were predisposed to CAD [OR 1.55, 95% CI (1.11, 2.16); P = 0.01]. Despite the functional evidence for a role of PARL in regulating insulin levels, no association with rs3732581 was found in the current study. Additionally, there were no associations with glucose levels, BMI or MetS. There were significant effects of the variant on mean IMT and risk of CAD. A role for PARL in metabolic conditions cannot be excluded and more comprehensive genetic studies are warranted.

The objective of this study was to determine whether single nucleotide polymorphisms (SNPs) in the Interleukin-1 (IL-1) gene family are associated with central obesity and metabolic syndrome in a coronary heart disease population. The IL-1 alpha C-889T (rs1800587) and IL-1 beta +3954 (rs1143634) SNPs were studied in a Western Australian coronary heart disease (CHD) population (N = 556). Subjects who were TT homozygous at either SNP had larger waist circumference (IL-1 alpha: 1.8 cm greater, P = 0.04; IL-1 beta: 4 cm greater, P = 0.0004) compared with major allele homozygotes. Individuals with two copies of the IL-1 alpha:IL-1 beta T:T haplotype had greater waist circumference (4.7 cm greater, P = 0.0001) compared to other haplotypes. There was a significant interaction between the IL-1 beta SNP and BMI level on waist circumference (P = 0.01). When the cohort was stratified by median BMI, TT carriers for IL-1 beta with above median BMI had greater waist circumference (6.1 cm greater, P = 0.007) compared to baseline carriers, whilst no significant association was seen in the below median group. Similarly, when the cohort was stratified by median fibrinogen level (IL-1 alpha interaction P = 0.01; IL-1 beta interaction P = 0.04), TT carriers for both SNPs in the above median fibrinogen group had greater waist circumference (IL-1 alpha 2.7 cm greater, P = 0.007; IL-1 beta 3.3 cm greater, P = 0.003) compared with major allele homozygotes. This association was not seen in the below median group. Also, we found a trend of increased metabolic syndrome for IL-1 beta TT homozygotes (P = 0.07). In conclusion, our findings suggest that in a CHD population IL-1 gene polymorphisms may be involved in increased central obesity, and the genetic influences are more evident among patients who have a higher level of obesity or inflammatory markers.

To access publisher's full text version of this article click on the hyperlink at the bottom of the page We performed fine mapping of 39 established type 2 diabetes (T2D) loci in 27,206 cases and 57,574 controls of European ancestry. We identified 49 distinct association signals at these loci, including five mapping in or near KCNQ1. 'Credible sets' of the variants most likely to drive each distinct signal mapped predominantly to noncoding sequence, implying that association with T2D is mediated through gene regulation. Credible set variants were enriched for overlap with FOXA2 chromatin immunoprecipitation binding sites in human islet and liver cells, including at MTNR1B, where fine mapping implicated rs10830963 as driving T2D association. We confirmed that the T2D risk allele for this SNP increases FOXA2-bound enhancer activity in islet- and liver-derived cells. We observed allele-specific differences in NEUROD1 binding in islet-derived cells, consistent with evidence that the T2D risk allele increases islet MTNR1B expression. Our study demonstrates how integration of genetic and genomic information can define molecular mechanisms through which variants underlying association signals exert their effects on disease. cademy of Finland 77299 102318 10493 118065 123885 124243 129293 129680 136895 139635 211119 213506 251217 263836 Agence National de la Recherche Association de Langue Francaise pour l'Etude du Diabete et des Maladies Metaboliques Association Diabeete Risque Vasculaire Association Francaise des Diabetiques Association of Danish Pharmacies Augustinus Foundation Becket Foundation British Diabetes Association (BDA) Research British Heart Foundation Central Norway Health Authority Central Finland Hospital District Center for Inherited Disease Research (CIDR) City of Kuopio City of Leutkirch Copenhagen County Danish Centre for Evaluation and Health Technology Assessment Danish Council for Independent Research Danish Heart Foundation Danish Research Councils Deutsche Forschungsgemeinschaft ER 155/6-2 Diabetes Research Foundation Diabetes UK Doris Duke Charitable Foundation Erasmus Medical Center Erasmus University Estonian government SF0180142s08 European Commission ENGAGE HEALTH-F4-2007-201413 FP7-201413 FP7-245536 EXGENESIS LSHM-CT-2004-005272 FP6 LSHM_CT_2006_037197 LSHM-CT-2007-037273 C-Public Health 2004310 European Regional Development Fund Federal Ministry of Education and Research, Germany FKZ 01GI1128 FKZ 01EO1001 Federal Ministry of Health, Germany Finnish Diabetes Association Finnish Diabetes Research Foundation Finnish Foundation for Cardiovascular Research Finnish Medical Society Folkhalsan Research Foundation Foundation for Life and Health in Finland Foundation for Old Servants Fredrick och Ingrid Thuring Foundation French region of Nord-Pas-de-Calais (Contrat de Projets Etat-Region) German Center for Diabetes Research German Research Council GRK1041 German National Genome Research Network Groupe d'Etude des Maladies Metaboliques et Systemiques Health Care Centers in Vasa, Narpes and Korsholm, Finland Health Foundation Heinz Nixdorf Foundation Helmholtz Zentrum Munchen Helsinki University Central Hospital Research Foundation Hospital District of Southwest Finland Ib Henriksens Foundation IngaBritt and Arne Lundberg's Research Foundation 359 Karolinska Institutet Knut and Alice Wallenberg Foundation KAW 2009.0243 Kuopio University Hospital Lundbeck Foundation Magnus Bergvall Foundation Medical Faculty of University Duisburg-Essen Medical Research Council, UK G0000649 G0601261 Ministry for Health, Welfare and Sports, the Netherlands Ministry of Education and Culture, Finland 722 627 Ministry of Education, Culture and Science, the Netherlands Ministry of Health and Prevention, Denmark Ministry of Social Affairs and Health, Finland Ministry of Innovation, Science, Research and Technology of North Rhine-Westphalia, Germany Munich Center of Health Sciences Municipal Health Care Center and Hospital in Jakobstad, Finland municipality of Rotterdam, the Netherlands Narpes Health Care Foundation National Health Screening Service of Norway National Heart, Lung, and Blood Institute, USA HHSN268201100005C HHSN268201100006C HHSN268201100007C HHSN268201100008C HHSN268201100009C HHSN268201100010C HHSN268201100011C HHSN268201100012C N01HC25195 N02HL64278 R01HL087641 R01HL59367 R01HL086694 National Human Genome Research Institute, USA U01HG004402 N01HG65403 National Institute for Diabetes and Digestive and Kidney Diseases, USA R01DK078616 U01DK085526 K24DK080140 R01DK073490 National Institute for Health and Welfare, Finland National Institutes of Health, USA HHSN268200625226C UL1RR025005 R01DK062370 R01DK072193 1Z01HG000024 AG028555 AG08724 AG04563 AG10175 AG08861 U01HG004399 DK58845 CA055075 DK085545 DK098032 Netherlands Genomics Initiative Netherlands Organisation for Health Research and Development Netherlands Organisation of Scientific Research NOW Investments 175.010.2005.011 911-03-012 050-060-810 Nord-Trondelag County Council Nordic Center of Excellence in Disease Genetics Norwegian Institute of Public Health Norwegian Research Council Novo Nordisk Foundation Ollquist Foundation Oxford National Institute for Health Research (NIHR) Biomedical Research Centre Paavo Nurmi Foundation Paivikki and Sakari Sohlberg Foundation Perklen Foundation Pirkanmaa Hospital District, Finland Programme Hospitalier de Recherche Clinique Programme National de Recherche sur la Diabete Research Institute for Diseases in the Elderly 014-93-015 Robert Dawson Evans Endowment, Department of Medicine, Boston University School of Medicine and Boston Medical Center Royal Swedish Academy of Sciences Sarstedt, Germany Signe and Ane Gyllenberg Foundation Sigrid Juselius Foundation Slottery Machine Association, Finland Social Insurance Institution of Finland South OstroBothnia Hospital District state of Baden-Wurttemberg, Germany Stockholm County Council 560183 Swedish Cultural Foundation, Finland Swedish Diabetes Foundation Swedish e-science Research Center Swedish Foundation for Strategic Research Swedish Heart-Lung Foundation Swedish Research Council SFO EXODIAB 2009-1039 521-2010-3490 521-2007-4037 521-2008-2974 ANDIS 825-2010-5983 LUDC 349-2008-6589 8691 Swedish Society of Medicine Tore Nilsson Foundation Torsten and Ragnar Soderbergs Stiftelser MT33/09 University Hospital Essen University of Tromso University College London NIHR Biomedical Research Centre UK NIHR Cambridge Biomedical Research Centre Uppsala University Uppsala University Hospital Vaasa Hospital District Velux Foundation Wellcome Trust GR072960 076113 083948 090367 090532 083270 086596 098017 095101 098051 098381

Genome-wide association scans are rapidly becoming reality, but there is no present consensus regarding genotyping strategies to optimise the discovery of true genetic risk factors. For a given investment in genotyping, should tag SNPs be selected in a gene-centric manner, or instead, should coverage be optimised based on linkage disequilibrium alone? We explored this question using empirical data from the HapMap-ENCODE project, and we found that tags designed specifically to capture common variation in exonic and evolutionarily conserved regions provide good coverage for 15-30% of the total common variation (depending on the population sample studied), and yield genotype savings compared with an anonymous tagging approach that captures all common variation. However, the same number of tags based on linkage disequilibrium alone captures substantially more (30-46%) of the total common variation. Therefore, the best strategy depends crucially on the unknown degree to which functional variation resides in recognisable exons and evolutionarily conserved sequence. A hypothetical but reasonable scenario might be one in which trait-causing variation is equally distributed between exons plus conserved sequence, and the rest of the genome. In this scenario, our analysis suggests that a tagging approach that captures variation in exons and conserved sequence provides only modestly better coverage of putatively causal variation than does anonymous tagging. In HapMap CEU samples (with northern and western European ancestry), we observed roughly equivalent coverage for equal investment for both tagging strategies.

It is well established that gene interactions influence common human diseases, but to date linkage studies have been constrained to searching for single genes across the genome. We applied a novel approach to uncover significant gene-gene interactions in a systematic two-dimensional (2D) genome-scan of essential hypertension. The study cohort comprised 2076 affected sib-pairs and 66 affected half-sib-pairs of the British Genetics of HyperTension study. Extensive simulations were used to establish significance thresholds in the context of 2D genome-scans. Our analyses found significant and suggestive evidence for loci on chromosomes 5, 9,11, 15, 16 and 19, which influence hypertension when gene-gene interactions are taken into account (5q13.1 and 11q22.1, two-locus lod score = 5.72; 5q13.1 and 19q12, two-locus lod score = 5.35; 9q22.3 and 15q12, two-locus lod score = 4.80; 16p12.3 and 16q23.1, two-locus lod score= 4.50). For each significant and suggestive pairwise interaction, the two-locus genetic model that best fitted the data was determined. Regions that were not detected using single-locus linkage analysis were identified in the 2D scan as contributing significant epistatic effects. This approach has discovered novel loci for hypertension and offers a unique potential to use existing data to uncover novel regions involved in complex human diseases.

The insulin-degrading enzyme is responsible for the intracellular proteolysis of insulin. Its gene IDE is located on chromosome 10, in an area with suggestive linkage to type 2 diabetes and related phenotypes. Due to the impact of genetic variants of this gene in rodents and the function of its protein product, it has been proposed as a candidate gene for type 2 diabetes. Various groups have explored the role of the common genetic variation of IDE on insulin resistance and reported associations of various single nucleotide polymorphisms (SNPs) and haplotypes on both type 2 diabetes and glycemic traits. We sought to characterize the haplotype structure of IDE in detail and replicate the association of common variants with type 2 diabetes, fasting insulin, fasting glucose, and insulin resistance. We assessed linkage disequilibrium, selected single-marker and multimarker tags, and genotyped these markers in several case-control and family-based samples totalling 4,206 Caucasian individuals. We observed no statistically significant evidence of association between single-marker or multimarker tests in IDE and type 2 diabetes. Nominally significant differences in quantitative traits are consistent with statistical noise. We conclude that common genetic variation at IDE is unlikely to confer clinically significant risk of type 2 diabetes in Caucasians.

The evaluation of results from primary genomewide linkage scans of complex human traits remains an area of importance and considerable debate. Apart from the usual assessment of statistical significance by use of asymptotic and empirical calculations, an additional means of evaluation--based on counting the number of distinct regions showing evidence of linkage--is possible. We have explored the characteristics of such a locus-counting method over a range of experimental conditions typically encountered during genomewide scans for complex trait loci. Under the null hypothesis, factors that have an impact on the informativeness of the data--such as map density, availability of parental data, and completeness of genotyping--are seen to markedly influence the number of regions of excess allele sharing and the empirically derived genomewide significance of the associated LOD score thresholds. In some circumstances, the expected number of regions is less than one-quarter of that predicted under the assumption of a dense map and complete extraction of inheritance information. We have applied this method to a previously analyzed data set--the Warren 2 genome scan for type 2-diabetes susceptibility--and demonstrate that more regions showing evidence for linkage were observed in the primary genome scan than would be expected by chance, across the whole range of LOD scores, even though no single linkage result achieved empirical genomewide statistical significance. Locus counting may be useful in assessing the results from genome scans for complex traits in general, especially because relatively few scans generate evidence for linkage reaching genomewide significance by dense-map criteria. By taking account of the effects of reduced data informativeness on the expected number of regions showing evidence for linkage, a more meaningful, and less conservative, evaluation of the results from such linkage studies is possible.

Immunohistochemistry is a method that can provide complementary diagnostic and prognostic information to morphological observations and soluble assays. Sensitivity, specificity, or requirements for arduous sample preparation or signal amplification procedures often limit the application of this approach to routine clinical specimens. Rolling circle amplification (RCA) generates a localized signal via an isothermal amplification of an oligonucleotide circle. The application of this approach to immunohistochemistry could extend the utility of these methods to include a more complete set of immunological and molecular probes. RCA-mediated signal amplification was successfully applied to the sensitive and specific detection of a variety of cell surface antigens (CD3, CD20, and epithelial membrane antigen) and intracellular molecules (vimentin and prostate-specific antigen) within a variety of routinely fixed specimens, as well as samples prepared for flow cytometry. RCA technology, which has an intrinsically wide dynamic range, is a robust and simple procedure that can provide a universal platform for the localization of a wide variety of molecules as a function of either antigenicity or nucleic acid sequence. The use of RCA in this way could enhance the use of markers of current interest as well as permit the integration of emerging information from genomics and proteomics intocell- and tissue-based analyses.

Atopy describes a syndrome of immunoglobulin E (IgE)-mediated allergy that underlies asthma and infantile eczema. We have previously identified a locus on chromosome 13q14 that is linked to atopy and to the total serum immunoglobulin A concentration. We have therefore made a saturation genetic map of the region by typing 59 polymorphic microsatellite loci on chromosome 13q. Multipoint linkage analysis identified a 1-LOD support unit for the location of the atopy locus with a 7.5-cM region flanked by the loci D13S328 and D13S1269. The peak of linkage was at locus D13S161 with a nonparametric -log of P score of approximately 4.5. Parent of origin effects were present, with linkage primarily observed to paternally derived alleles. The genetic map of this region provides a basis for the effective identification of the chromosome 13 atopy gene.

We describe an adaptation of the rolling circle amplification (RCA) reporter system for the detection of protein Ags, termed “immunoRCA.” In immunoRCA, an oligonucleotide primer is covalently attached to an Ab; thus, in the presence of circular DNA, DNA polymerase, and nucleotides, amplification results in a long DNA molecule containing hundreds of copies of the circular DNA sequence that remain attached to the Ab and that can be detected in a variety of ways. Using immunoRCA, analytes were detected at sensitivities exceeding those of conventional enzyme immunoassays in ELISA and microparticle formats. The signal amplification afforded by immunoRCA also enabled immunoassays to be carried out in microspot and microarray formats with exquisite sensitivity. When Ags are present at concentrations down to fM levels, specifically bound Abs can be scored by counting discrete fluorescent signals arising from individual Ag–Ab complexes. Multiplex immunoRCA also was demonstrated by accurately quantifying Ags mixed in different ratios in a two-color, single-molecule-counting assay on a glass slide. ImmunoRCA thus combines high sensitivity and a very wide dynamic range with an unprecedented capability for single molecule detection. This Ag-detection method is of general applicability and is extendable to multiplexed immunoassays that employ a battery of different Abs, each labeled with a unique oligonucleotide primer, that can be discriminated by a color-coded visualization system. ImmunoRCA-profiling based on the simultaneous quantitation of multiple Ags should expand the power of immunoassays by exploiting the increased information content of ratio-based expression analysis.

Interpretation of dense single nucleotide polymorphism (SNP) follow-up of genome-wide association or linkage scan signals can be facilitated by establishing expectation for the behaviour of primary mapping signals upon fine-mapping, under both null and alternative hypotheses. We examined the inferences that can be made regarding the posterior probability of a real genetic effect and considered different disease-mapping strategies and prior probabilities of association. We investigated the impact of the extent of linkage disequilibrium between the disease SNP and the primary analysis signal and the extent to which the disease gene can be physically localised under these scenarios. We found that large increases in significance (>2 orders of magnitude) appear in the exclusive domain of genuine genetic effects, especially in the follow-up of genome-wide association scans or consensus regions from multiple linkage scans. Fine-mapping significant association signals that reside directly under linkage peaks yield little improvement in an already high posterior probability of a real effect. Following fine-mapping, those signals that increase in significance also demonstrate improved localisation. We found local linkage disequiliptium patterns around the primary analysis signal(s) and tagging efficacy of typed markers to play an important role in determining a suitable interval for fine-mapping. Our findings help inform the interpretation and design of dense SNP-mapping follow-up studies, thus facilitating discrimination between a genuine genetic effect and chance fluctuation (false positive).

Endothelin-1 is a potent vasoconstrictor in the body. Previous studies have identified associations between the coding polymorphism K198N and hypertension, systolic blood pressure and HDL levels. We sought to examine the evidence for these associations and, additionally, the association between K198N, insulin resistance, metabolic syndrome and coronary artery disease (CAD). We used generalised linear modelling to test K198N for association with hypertension and systolic blood pressure, lipid levels, insulin resistance scores and metabolic syndrome in a general cross-sectional community sample. Mean carotid intima media thickness and risk of carotid plaque were examined in the general population sample, and Gensini score was examined in a sample of patients with CAD. A case/control sample was used to examine the association of K198N with risk of CAD. There was no significant evidence for association between K198N and hypertension, systolic blood pressure, lipid levels, insulin resistance or metabolic syndrome in either population. The minor allele was marginally associated with increased mean IMT levels (P = 0.02) in the general population sample, although not with CAD in the case/control study or with the severity of disease in patients with CAD. In conclusion, we found no robust evidence for the associations between K198N and hypertension, systolic blood pressure or HDL levels seen in previous studies.

Serum high density lipoprotein (HDL) levels are inversely related to the development of coronary artery disease (CAD). Apolipoproteins AI and AII are the major protein constituents of HDL particles. APOAI and APOAII genetic polymorphisms have been proposed to affect transcriptional efficiency of their respective genes, thereby altering serum lipid levels and influencing atherosclerotic disease risk. 556 subjects with angiographically proven CAD (>50% stenosis) and 1109 randomly selected individuals from metropolitan Perth, Western Australia, were included in an association study. APOAI -75G/A (rs670) and APOAII -256T/C (rs5082) polymorphisms were both found to be not associated with plasma HDL levels. In a case-control analysis of 484 male CAD patients and 498 male controls, individuals carrying the 'CC' genotype for the APOAII rs5082 polymorphism had significantly lower risk of CAD than the 'T' allele carriers (OR=0.57, 95% CI 0.39-0.84, p=0.004). The minor 'A' allele of the APOAI rs670 polymorphism was found to be not associated with CAD, contrary to previous reports. We conclude that the APOAII rs5082 polymorphism appears to be cardioprotective in this representative Caucasian Australian population.

CONTEXT: Recently, a quantitative trait locus for stature was reported on chromosome 3p26 in patients with type 2 diabetes. OBJECTIVE: Given that ghrelin is a peptide involved in GH release and located on 3p26, we hypothesized that variation within its gene (GHRL) may be responsible for the quantitative trait locus on 3p26. DESIGN: The evidence for linkage around GHRL was refined with the genotyping of an additional four microsatellites (D3S4545, D3S1537, D3S1597, and D3S3611), giving a total of 27 markers, followed by multipoint variance components linkage analysis. Probands from the linkage families were typed for five common single nucleotide polymorphisms (SNPs) within GHRL and tested for association with adult stature using haplotype trend regression. RESULTS: The maximum multipoint evidence for linkage between adult stature and the 27 microsatellites yielded an LOD score of 2.58 (P = 0.0003) between D3S1297 and D3S1304. Five common (frequency of > or =5%) SNPs were typed in the probands [two promoter SNPs (rs27647 and rs26802), two exonic (rs696217 and rs4684677), and one intronic (rs35683)] capturing 80% of the total common variation in GHRL. No association was found between any SNP (or haplotypes thereof) and adult stature. CONCLUSION: Common genetic variation within GHRL is not responsible for variation in adult stature in this population.

Characterisation of the interactions between susceptibility loci (epistasis) is central to a full understanding of the genetic aetiology and the molecular pathology of complex diseases. We have examined, in British and French pedigrees, evidence for epistasis between the type 2 diabetes susceptibility loci on chromosomes 1q21-25 and 10q23-26 using two complementary linkage-based approaches. Joint two-locus linkage analysis of 1q and 10q in British pedigrees provided significant evidence for interaction (P < or = 0.003) when comparing a general epistasis model with multiplicative or additive-effects-only models. Conditional linkage analysis (which models epistasis as a deviation from multiplicativity only) confirmed these findings, with significant LOD score increases at the 1q (P = 0.0002) and 10q (P = 0.0023) loci. These analyses provided sizeable reductions in the 1-LOD support intervals for both loci. Analyses of the British and French pedigrees together yielded comparable, but not enhanced, findings, with significant (P < or = 0.003) evidence for epistasis in joint two-locus linkage analysis, and during conditional linkage analysis significant increases in linkage evidence at the 1q (P = 0.0002) and 10q (P = 0.0036) loci. Our findings of epistasis nevertheless substantiate the evidence for genuine genetic effects at both loci, facilitate endeavours to fine-map these loci in population samples, and support further examination of this interaction at the nucleotide level by providing a robust prior hypothesis.

Recent data suggest that common variation in the transcription factor 7-like 2 ( TCF7L2 ) gene is associated with type 2 diabetes. Evaluation of such associations in independent samples provides necessary replication and a robust assessment of effect size. Using four TCF7L2 single nucleotide polymorphisms (SNPs; including the two most associated in the previous study), we conducted a case-control study in 2,158 type 2 diabetic subjects and 2,574 control subjects and a family-based association analysis in 388 parent-offspring trios all from the U.K. All SNPs showed powerful associations with diabetes in the case-control analysis, with strongest effects at rs7903146 (allele-wise relative risk 1.36 [95% CI 1.24–1.48], P = 1.3 × 10 −11 ). Data were consistent with a multiplicative model. The family-based analyses provided independent evidence for association at all loci (e.g., rs4506565, 62% transmission, P = 7 × 10 −5 ) with no parent-of-origin effects. The frequency of diabetes-associated TCF7L2 genotypes was greater in cases ascertained for positive family history and early onset (rs4606565, P = 0.02); the population-attributable risk, estimated from the least-selected cases, is ∼16%. The overall evidence for association for these variants ( P = 4.4 × 10 −14 combining case-control and family-based analyses for rs4506565) exceeds genome-wide significance criteria and clearly establishes TCF7L2 as a type 2 diabetes susceptibility gene of substantial importance.

Obesity is a major health problem, and many family-based studies have suggested that it has a strong genetic basis. We performed a genome-wide quantitative trait linkage scan for loci influencing BMI in 573 pedigrees from the U.K. We identified genome-wide significant linkage (logarithm of odds = 3.74, between D10S208 and D10S196, genome-wide P = 0.0186) on chromosome 10p. The size of our study population and the statistical significance of our findings provide substantial contributions to the body of evidence for a locus on chromosome 10p. We examined eight single nucleotide polymorphisms (SNPs) in GAD2, which maps to this linkage region, tagging the majority of variation in the gene, and observed marginally significant (0.01 < P < 0.05) associations between four common variants and BMI. However, these SNPs did not account for our evidence of linkage to BMI, and they did not replicate (in direction of effect) the previous associations. We therefore conclude that these SNPs are not the etiological variants underlying this locus. We cannot rule out the possibility that other untagged variations in GAD2 may, in part, be involved, but it is most likely that alternative gene(s) within the broad gene-rich region of linkage on 10p are responsible for variation in body mass and susceptibility to obesity.

The insulin-degrading enzyme is responsible for the intracellular proteolysis of insulin. Its gene IDE is located on chromosome 10, in an area with suggestive linkage to type 2 diabetes and related phenotypes. Due to the impact of genetic variants of this gene in rodents and the function of its protein product, it has been proposed as a candidate gene for type 2 diabetes. Various groups have explored the role of the common genetic variation of IDE on insulin resistance and reported associations of various single nucleotide polymorphisms (SNPs) and haplotypes on both type 2 diabetes and glycemic traits. We sought to characterize the haplotype structure of IDE in detail and replicate the association of common variants with type 2 diabetes, fasting insulin, fasting glucose, and insulin resistance. We assessed linkage disequilibrium, selected single-marker and multimarker tags, and genotyped these markers in several case-control and family-based samples totalling 4,206 Caucasian individuals. We observed no statistically significant evidence of association between single-marker or multimarker tests in IDE and type 2 diabetes. Nominally significant differences in quantitative traits are consistent with statistical noise. We conclude that common genetic variation at IDE is unlikely to confer clinically significant risk of type 2 diabetes in Caucasians.

The two-stage linkage mapping protocol for complex traits (a primary genome scan with low marker density followed by the high-density genotyping around linkage peaks) is a near-universal practice. The behavior (an increase or a decrease) of the peak upon such fine mapping frequently leads to inferences regarding the veracity of the primary scan finding, namely a true, or a false, positive. We examined by simulation, under the null hypothesis of no linkage and the alternative hypothesis of true linkage, the inferences that can be made regarding the posterior probability of linkage given either a peak increase, or alternatively, a peak decrease, following fine mapping. We considered different models of missing genotype data, fine-mapping LOD score thresholds, and prior probabilities of linkage. Our simulations show that evidence for linkage can increase frequently upon fine mapping under both null and alternative hypotheses, although large increases in LOD scores are more common under the alternative hypothesis. Increased LOD scores accompany an increased posterior probability of linkage, and large LOD score changes and the presence of dominance at the trait locus accentuate this effect. We demonstrate that the greatest changes in the posterior probability of linkage occur when the genotyping data are least complete (and especially when parental genotypes are missing), and the LOD score threshold for fine mapping is relaxed. Genet. Epidemiol. © 2004 Wiley-Liss, Inc.