A role for coding functional variants in HNF4A in type 2 diabetes susceptibility.

AIMS/HYPOTHESIS: Rare mutations in the gene HNF4A, encoding the transcription factor hepatocyte nuclear factor 4α (HNF-4A), account for ~5% of cases of MODY and more frequent variants in this gene may be involved in multifactorial forms of diabetes. Two low-frequency, non-synonymous variants in HNF4A (V255M, minor allele frequency [MAF] ~0.1%; T130I, MAF ~3.0%)-known to influence downstream HNF-4A target gene expression-are of interest, but previous type 2 diabetes association reports were inconclusive. We aimed to evaluate the contribution of these variants to type 2 diabetes susceptibility through large-scale association analysis. METHODS: We genotyped both variants in at least 5,745 cases and 14,756 population controls from the UK and Denmark. We also undertook an expanded association analysis that included previously reported and novel genotype data obtained in Danish, Finnish, Canadian and Swedish samples. A meta-analysis incorporating all published association studies of the T130I variant was subsequently carried out in a maximum sample size of 14,279 cases and 26,835 controls. RESULTS: We found no association between V255M and type 2 diabetes in either the initial (p = 0.28) or the expanded analysis (p = 0.44). However, T130I demonstrated a modest association with type 2 diabetes in the UK and Danish samples (additive per allele OR 1.17 [95% CI 1.08-1.28]; p = 1.5 × 10⁻4), which was strengthened in the meta-analysis (OR 1.20 [95% CI 1.10-1.30]; p = 2.1 × 10⁻5). CONCLUSIONS/INTERPRETATION: Our data are consistent with T130I as a low-frequency variant influencing type 2 diabetes risk, but are not conclusive when judged against stringent standards for genome-wide significance. This study exemplifies the difficulties encountered in association testing of low-frequency variants.

Lod scores for gene mapping in the presence of marker map uncertainty.

Multipoint lod scores are typically calculated for a grid of locus positions, moving the putative disease locus across a fixed map of genetic markers. Changing the order of a set of markers and/or the distances between the markers can make a substantial difference in the resulting lod score curve and the location and height of its maximum. The typical approach of using the best maximum likelihood marker map is not easily justified if other marker orders are nearly as likely and give substantially different lod score curves. To deal with this problem, we propose three weighted multipoint lod score statistics that make use of information from all plausible marker orders. In each of these statistics, the information conditional on a particular marker order is included in a weighted sum, with weight equal to the posterior probability of that order. We evaluate the type 1 error rate and power of these three statistics on the basis of results from simulated data, and compare these results to those obtained using the best maximum likelihood map and the map with the true marker order. We find that the lod score based on a weighted sum of maximum likelihoods improves on using only the best maximum likelihood map, having a type 1 error rate and power closest to that of using the true marker order in the simulation scenarios we considered.

The Finland-United States investigation of non-insulin-dependent diabetes mellitus genetics (FUSION) study. II. An autosomal genome scan for diabetes-related quantitative-trait loci.

Type 2 diabetes mellitus is a complex disorder encompassing multiple metabolic defects. We report results from an autosomal genome scan for type 2 diabetes-related quantitative traits in 580 Finnish families ascertained for an affected sibling pair and analyzed by the variance components-based quantitative-trait locus (QTL) linkage approach. We analyzed diabetic and nondiabetic subjects separately, because of the possible impact of disease on the traits of interest. In diabetic individuals, our strongest results were observed on chromosomes 3 (fasting C-peptide/glucose: maximum LOD score [MLS] = 3.13 at 53.0 cM) and 13 (body-mass index: MLS = 3.28 at 5.0 cM). In nondiabetic individuals, the strongest results were observed on chromosomes 10 (acute insulin response: MLS = 3.11 at 21.0 cM), 13 (2-h insulin: MLS = 2.86 at 65.5 cM), and 17 (fasting insulin/glucose ratio: MLS = 3.20 at 9.0 cM). In several cases, there was evidence for overlapping signals between diabetic and nondiabetic individuals; therefore we performed joint analyses. In these joint analyses, we observed strong signals for chromosomes 3 (body-mass index: MLS = 3.43 at 59.5 cM), 17 (empirical insulin-resistance index: MLS = 3.61 at 0.0 cM), and 19 (empirical insulin-resistance index: MLS = 2.80 at 74.5 cM). Integrating genome-scan results from the companion article by Ghosh et al., we identify several regions that may harbor susceptibility genes for type 2 diabetes in the Finnish population.

Point and interval estimates of marker location in radiation hybrid mapping.

Radiation hybrid (RH) mapping is a powerful method for ordering loci on chromosomes and for estimating the distances between them. RH mapping is currently used to construct both framework maps, in which all markers are ordered with high confidence (e.g., 1,000:1 relative maximum likelihood), and comprehensive maps, which include markers with less-confident placement. To deal with uncertainty in the order and location of markers, marker positions may be estimated conditional on the most likely marker order, plausible intervals for nonframework markers may be indicated on a framework map, or bins of markers may be constructed. We propose a statistical method for estimating marker position that combines information from all plausible marker orders, gives a measure of uncertainty in location for each marker, and provides an alternative to the current practice of binning. Assuming that the prior distribution for the retention probabilities is uniform and that the marker loci are distributed independently and uniformly on an interval of specified length, we calculate the posterior distribution of marker position for each marker. The median or mean of this distribution provides a point estimate of marker location. An interval estimate of marker location may be constructed either by using the 100(alpha/2) and 100(1-alpha)/2 percentiles of the distribution to form a 100(1-alpha) % posterior credible interval or by calculating the shortest 100(1-alpha) % posterior credible interval. These point and interval estimates take into account ordering uncertainty and do not depend on the assumption of a particular marker order. We evaluate the performance of the estimates on the basis of results from simulated data and illustrate the method with two examples.

Autosomal dominant nanophthalmos (NNO1) with high hyperopia and angle-closure glaucoma maps to chromosome 11.

Nanophthalmos is an uncommon developmental ocular disorder characterized by a small eye, as indicated by short axial length, high hyperopia (severe farsightedness), high lens/eye volume ratio, and a high incidence of angle-closure glaucoma. We performed clinical and genetic evaluations of members of a large family in which nanophthalmos is transmitted in an autosomal dominant manner. Ocular examinations of 22 affected family members revealed high hyperopia (range +7.25-+13.00 diopters; mean +9.88 diopters) and short axial length (range 17.55-19.28 mm; mean 18.13 mm). Twelve affected family members had angle-closure glaucoma or occludable anterior-chamber angles. Linkage analysis of a genome scan demonstrated highly significant evidence that nanophthalmos in this family is the result of a defect in a previously unidentified locus (NNO1) on chromosome 11. The gene was localized to a 14.7-cM interval between D11S905 and D11S987, with a maximum LOD score of 5. 92 at a recombination fraction of .00 for marker D11S903 and a multipoint maximum LOD score of 6.31 for marker D11S1313. NNO1 is the first human locus associated with nanophthalmos or with an angle-closure glaucoma phenotype, and the identification of the NNO1 locus is the first step toward the cloning of the gene. A cloned copy of the gene will enable examination of the relationship, if any, between nanophthalmos and less severe forms of hyperopia and between nanophthalmos and other conditions in which angle-closure glaucoma is a feature.

Novel trabecular meshwork inducible glucocorticoid response mutation in an eight-generation juvenile-onset primary open-angle glaucoma pedigree.

OBJECTIVE: This study aimed to update a large kindred with juvenile-onset primary open-angle glaucoma (POAG) first described in 1940 and to identify the underlying genetic cause of the disease. DESIGN: Molecular genetic study of a single kindred, including clinical examination, retrospective review of clinical and family history records, linkage analysis, and mutation screening. PARTICIPANTS: The retrospective review included 957 members of a single large family. The linkage study included 40 members of 1 branch of the family in which juvenile-onset POAG is segregating in an autosomal-dominant pattern. Mutation screening included 15 at-risk family members with juvenile-onset POAG, probands of 40 families with adult-onset POAG, probands of 11 additional unrelated juvenile-onset POAG families, and 43 unrelated normal control subjects. INTERVENTION: Clinical and family history records were obtained, ophthalmologic examinations were performed, and blood samples were drawn for use in genotyping. MAIN OUTCOME MEASURES: Allele sizes of microsatellite repeat genetic markers from the vicinity of the GLC1A glaucoma gene on chromosome 1q were assigned based on size fractionation of DNA fragments generated by polymerase chain reaction (PCR). Linkage was established by the method of lod scores. Mutations were identified by determination of the DNA sequence of PCR products amplified from the trabecular meshwork inducible glucocorticoid response (TIGR) gene. Glaucoma status for purposes of linkage and mutation analysis was based on a combination of ophthalmologic examination, clinical records, family history, and previously published information. For some individuals reported in the pedigree, but not included in the genotyping studies, less information was available as presented in the text and tables. RESULTS: Autosomal-dominant POAG was confirmed or reported for 78 members of an 8-generation family. Linkage analysis showed significant evidence for linkage of juvenile-onset POAG in one branch of the family to D1S452 (maximum lod score of 6.42 at a recombination fraction of 0.00) and other markers in the vicinity of the GLC1A gene on chromosome 1q. Screening of the TIGR gene identified a mutation that results in substitution of asparagine for isoleucine at codon 477 near the carboxyterminal end of the protein. CONCLUSIONS: The authors' findings strongly suggest that the juvenile-onset POAG locus in this family is the GLC1A locus and that the underlying cause of the disease is the IIe477Asn TIGR mutation that cosegregates with juvenile-onset POAG in one branch of this large family. Lack of samples from deceased individuals prevented the authors from determining whether reported adult-onset cases in this family could also be attributed to the IIe477Asn TIGR mutation. Absence of the IIe477Asn TIGR mutation from other juvenile- and adult-onset POAG families implies that this TIGR mutation is not a common cause of glaucoma.

Fine localization of the Nijmegen breakage syndrome gene to 8q21: evidence for a common founder haplotype.

Nijmegen breakage syndrome (NBS) is a rare autosomal recessive disorder characterized by microcephaly, a birdlike face, growth retardation, immunodeficiency, lack of secondary sex characteristics in females, and increased incidence of lymphoid cancers. NBS cells display a phenotype similar to that of cells from ataxia-telangiectasia patients, including chromosomal instability, radiation sensitivity, and aberrant cell-cycle-checkpoint control following exposure to ionizing radiation. A recent study reported genetic linkage of NBS to human chromosome 8q21, with strong linkage disequilibrium detected at marker D8S1811 in eastern European NBS families. We collected a geographically diverse group of NBS families and tested them for linkage, using an expanded panel of markers at 8q21. In this article, we report linkage of NBS to 8q21 in 6/7 of these families, with a maximum LOD score of 3.58. Significant linkage disequilibrium was detected for 8/13 markers tested in the 8q21 region, including D8S1811. In order to further localize the gene for NBS, we generated a radiation-hybrid map of markers at 8q21 and constructed haplotypes based on this map. Examination of disease haplotypes segregating in 11 NBS pedigrees revealed recombination events that place the NBS gene between D8S1757 and D8S270. A common founder haplotype was present on 15/18 disease chromosomes from 9/11 NBS families. Inferred (ancestral) recombination events involving this common haplotype suggest that NBS can be localized further, to an interval flanked by markers D8S273 and D8S88.

Cosegregation of open-angle glaucoma and the nail-patella syndrome.

PURPOSE: To evaluate two families ascertained only for the presence of glaucoma in which both nail-patella syndrome and glaucoma occur in several generations and to determine whether the two diseases are genetically related. METHODS: Ophthalmologic examinations and orthopedic examinations were performed. DNA samples from family members were screened with a microsatellite repeat marker at the argininosuccinate synthetase (ASS) locus at 9q34, and linkage analysis was performed. RESULTS: Six patients with open-angle glaucoma were found among 13 patients with nail-patella syndrome in family UM:47. Seven patients with glaucoma were found among 11 patients with nail-patella syndrome in family UM:65. In both families, all individuals with glaucoma also had nail-patella syndrome. Two-point linkage analysis resulted in a lod score of 2.98 at a recombination fraction of 0.00 for open-angle glaucoma and nail-patella syndrome. CONCLUSIONS: Linkage results presented here provide strong evidence that the orthopedic and nail anomalies in these two families result from the same nail-patella syndrome locus that has been previously linked to markers at 9q34. These data provide indirect evidence for a possible glaucoma locus at 9q34 and do not allow us to distinguish whether the glaucoma is the result of the nail-patella syndrome mutation or whether there is a separate locus responsible for glaucoma in these families. These studies suggest a need for ophthalmologic examination of individuals with nail-patella syndrome.

Juvenile glaucoma linked to the GLC1A gene on chromosome 1q in a Panamanian family.

PURPOSE: To characterize clinically and genetically autosomal dominant juvenile-onset primary open-angle glaucoma in a Panamanian family. METHODS: Twenty members of a six-generation family underwent ophthalmologic examination and genetic screening with markers near the GLC1A gene on chromosome 1q. RESULTS: Linkage analysis disclosed evidence linking primary open-angle glaucoma in this family to the GLC1A gene on chromosome 1q, with a maximum lod score of 3.75 for marker D1S431 at an estimated recombination fraction of 0.00. CONCLUSIONS: This is the first report of a Panamanian family in which primary open-angle glaucoma is linked to the GLC1A gene on chromosome 1q.

Identifying marker typing incompatibilities in linkage analysis.

A common problem encountered in linkage analyses is that execution of the computer program is halted because of genotypes in the data that are inconsistent with Mendelian inheritance. Such inconsistencies may arise because of pedigree errors or errors in typing. In some cases, the source of the inconsistencies is easily identified by examining the pedigree. In others, the error is not obvious, and substantial time and effort are required to identify the responsible genotypes. We have developed two methods for automatically identifying those individuals whose genotypes are most likely the cause of the inconsistencies. First, we calculate the posterior probability of genotyping error for each member of the pedigree, given the marker data on all pedigree members and allowing anyone in the pedigree to have an error. Second, we identify those individuals whose genotypes could be solely responsible for the inconsistency in the pedigree. We illustrate these methods with two examples: one a pedigree error, the second a genotyping error. These methods have been implemented as a module of the pedigree analysis program package MENDEL.

Clinical phenotype of juvenile-onset primary open-angle glaucoma linked to chromosome 1q.

PURPOSE: Recent reports have suggested that a gene responsible for juvenile-onset primary open-angle glaucoma exists on the long arm of chromosome 1 (1q). This report describes a previously unpublished family (UM:JG3) in which juvenile-onset glaucoma is segregating in an autosomal dominant manner. The clinical features in this family were compared with those seen in other pedigrees with this condition. Linkage analysis was performed to evaluate whether a glaucoma-causing gene in UM:JG3 is linked to genetic markers on chromosome 1q. METHODS: Affected family members, their siblings, children, and spouses were examined to identify the presence of glaucoma. Linkage studies were performed using short tandem repeat polymorphisms from chromosome 1q. Results of these studies were compared with those found for other families in which juvenile-onset primary open-angle glaucoma is linked genetically to the same chromosome 1q region. RESULTS: The UM:JG3 family includes 22 affected individuals over five generations, including 12 still living. The average age at diagnosis for living affected individuals was 26 years. An association between myopia and glaucoma was observed in this family, but the glaucoma was not associated with iris processes or other structural anomalies. The clinical course of disease and response to treatment were similar to other families with this disease. The disease phenotype in this family is linked to markers on chromosome 1q with a maximum lod score of 3.52 at a recombination fraction of 0.00 for marker D1S433. Haplotype analysis suggests the gene responsible for glaucoma in this family is located in an 8-cM region between markers D1S445 and D1S218. CONCLUSIONS: The glaucoma in UM:JG3 is linked to markers on chromosome 1q, with a candidate interval smaller than that in previous reports. In individuals with juvenile-onset open-angle glaucoma linked to chromosome 1q, the phenotype can range from mild ocular hypertension to blindness, resulting from marked elevations in intraocular pressure, with age at diagnosis ranging from 6 to 62 years. However, most affected individuals display a characteristic phenotype that includes onset in the first three decades of life, unusually high intraocular pressures, and the need for surgical therapy to prevent loss of vision. Whether differences in expression among families is due to allelic heterogeneity remains to be determined.

Juvenile glaucoma linked to GLCIA in a Panamanian family.

PURPOSE: To carry out clinical and genetic characterization of juvenile-onset primary open-angle glaucoma (POAG) inherited as an autosomal dominant trait in a Panamanian family. METHODS: Twenty-two members of a six-generation Panamanian family underwent an ophthalmologic evaluation. Blood samples were collected from 20 of these individuals for preparation of DNA for use in screening of microsatellite repeat genetic markers via polymerase chain reaction. RESULTS: Eleven living family members covering 4 generations were diagnosed as affected with open-angle glaucoma of primarily juvenile onset. Four of 6 other at-risk individuals examined and enrolled were characterized as unaffected and two as indeterminate. Two additional individuals were not included in this study because they were too young to characterize or to provide a blood sample. Three spouses of affected family members were also examined and found not to have glaucoma. Of clinical importance was the finding of markedly elevated intraocular pressure (IOP) in 2 affected brothers, both of whom were advised to have urgent filtration surgery; the finding of elevated IOP in the only seeing eye of the mother of these brothers, causing us to advise her to pursue more aggressive treatment; and the finding of early signs of glaucoma in a previously undiagnosed 9-year-old family member. Linkage analysis using selected microsatellite repeat markers in the 1q21-q31 region revealed strong evidence for linkage to the GLC1A gene with a maximum lod score of 3.75 for marker D1S431 at a recombination fraction of 0.00. CONCLUSIONS: The most likely interpretation of our data is that a mutation in the GLC1A gene is responsible for juvenile-onset POAG in this Panamanian family, thus expanding the countries of origin where this gene has been found to exist. The numbers of families with GLC1A glaucoma now reported from only a few centers worldwide raise questions about whether this disease may be more common than once thought. Evaluation of treatment histories and clinical outcomes in members of this and other previously reported families indicates that ophthalmologists need to understand the necessity for urgent filtration surgery in most cases of GLC1A glaucoma if vision is to be preserved.