Construction of a 1-Mb restriction-mapped cosmid contig containing the candidate region for the familial mediterranean fever locus (MEFV) on chromosome 16p13.3

In this paper we describe the assembly and restriction map of a 1.05-Mb cosmid contig spanning the candidate region for familial Mediterranean fever (FMF), a recessively inherited disorder of inflammation localized to 16p13.3. Using a combination of cosmid walking and screening for P1, PAC, BAG, and YAC clones, we have generated a contig of genomic clones spanning similar to 1050 kb that contains the FMF critical region. The map consists of 179 cosmid, 15 P1, 10 PAC, 3 BAG, and 17 YAC clones, anchored by 27 STS markers. Eight additional STSs have been developed from the similar to 700 kb immediately centromeric to this genomic region. Five of the 35 STSs are microsatellites that have not been previously reported. NotI and EcoRI mapping of the overlapping cosmids, hybridization of restriction fragments from cosmids to one another, and STS analyses have been used to validate the assembly of the contig. Our contig totally subsumes the 250-kb interval recently reported, by founder haplotype analysis, to contain the FMF gene. Thus, our high-resolution clone map provides an ideal resource for transcriptional mapping toward the eventual identification of this disease gene. (C) 1997 Academic Press.

Association between ADAMTS13 polymorphisms and risk of cardiovascular events in chronic coronary disease

Introduction: Association between ADAMTS13 levels and cardiovascular events has been described recently. However, no genetic study of ADAMTS13 in coronary patients has been described. Materials and Methods: Based on related populations frequencies and functional studies, we tested three ADAMTS13 polymorphisms: C1342G (Q448E), C1852G (P618A) and C2699T (A900V) in a group of 560 patients enrolled in the Medical, Angioplasty, or Surgery Study II (MASS II), a randomized trial comparing treatments for patients with coronary artery disease (CAD) and preserved left ventricular function. The incidence of the 5-year end-points of death and death from cardiac causes, myocardial infarction, refractory angina requiring revascularization and cerebrovascular accident was determined for each polymorphim`s allele, genotype and haplotype. Risk was assessed with the use of logistic regression and Cox proportional-hazards model and multivariable adjustment was employed for possible confounders. Results: Clinical characteristics and received treatment of each genotype group were similar at baseline. In an adjusted model for cardiovascular risk variables, we were able to observe a significant association between ADAMTS13 900V variant and an increased risk of death (OR: 1,92 CI: 1,14-3,23, p = 0,015) or death from cardiac cause (OR: 2,67, CI: 1,59-4,49, p = 0,0009). No association between events and ADAMTS13 Q448E or P618A was observed. Conclusions: This first report studying the association between ADAMTS13 genotypes and cardiovascular events provides evidence for the association between ADAMTS13 900V variant and an increased risk of death in a population with multi-vessel CAD. (C) 2009 Elsevier Ltd. All rights reserved.

European ancestry and polymorphisms in DNA repair genes modify the risk of melanoma: A case-control study in a high UV index region in Brazil

Background: UV radiation is the major environmental factor related to development of cutaneous melanoma. Besides sun exposure and the influence of latitude, some host characteristics such as skin phototype and hair and eye color are also risk factors for melanoma. Polymorphisms in DNA repair genes could be good candidates for susceptibility genes, mainly in geographical regions exposed to high solar radiation. Objective: Evaluate the role of host characteristic.; and DNA repair polymorphism in melanoma risk in Brazil. Methods: We carried out a hospital-based case-control study in Brazil to evaluate the contribution of host factors and polymorphisms in DNA repair to melanoma risk. A total of 412 patients (202 with melanoma and 210 controls) were analyzed regarding host characteristics for melanoma risk as well as for 11 polymorphisms in DNA repair genes. Results: We found an association of host characteristics with melanoma development, such as eye and hair color, fair skin, history of pigmented lesions removed, sunburns in childhood and adolescence, and also European ancestry. Regarding DNA repair gene polymorphisms, we found protection for the XPG 1104 His/His genotype (OR 0.32; 95% CI 0.13-0.75), and increased risk for three polymorphisms in the XPC gene (PAT+; IV-6A and 939Gln), which represent a haplotype for XPC. Melanoma risk was higher in individuals carrying the complete XPC haplotype than each individual polymorphism (OR 3.64; 95% CI 1.77-7.48). Conclusions: Our data indicate that the host factors European ancestry and XPC polymorphisms contributed to melanoma risk in a region exposed to high sun radiation. (C) 2011 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

Ancient missense mutations in a new member of the RoRet gene family are likely to cause familial Mediterranean fever

Familial Mediterranean fever (FMF) is a recessively inherited disorder characterized by dramatic episodes of fever and serosal inflammation. This report describes the cloning of the gene likely to cause FMF from a 115-kb candidate interval on chromosome 16p. Three different missense mutations were identified in affected individuals, but not in normals. Haplotype and mutational analyses disclosed ancestral relationships among carrier chromosomes in populations that have been separated for centuries. The novel gene encodes a 3.7-kb transcript that is almost exclusively expressed in granulocytes. The predicted protein, pyrin, is a member of a family of nuclear factors homologous to the Ro52 autoantigen. The cloning of the FMF gene promises to shed light on the regulation of acute inflammatory responses.

Genetic heterogeneity in familial acute myelogenous leukemia: Evidence for a second locus at chromosome 16q21-23.2

The identification of genes responsible for the rare cases of familial leukemia may afford insight into the mechanism underlying the more common sporadic occurrences. Here we test a single family with 11 relevant meioses transmitting autosomal dominant acute myelogenous leukemia (AML) and myelodysplasia for linkage to three potential candidate loci. In a different family with inherited AML, linkage to chromosome 21q22.1-22.2 was recently reported; we exclude linkage to 21q22.1-22.2, demonstrating that familial AML is a heterogeneous disease. After reviewing familial leukemia and observing anticipation in the form of a declining age of onset with each generation, we had proposed 9p21-22 and 16q22 as additional candidate loci. Whereas linkage to 9p21-22 can be excluded, the finding of a maximum two-point LOD score of 2.82 with the microsatellite marker D16S522 at a recombination fraction theta = 0 provides evidence supporting linkage to 16q22. Haplotype analysis reveals a 23.5-cM (17.9-Mb) commonly inherited region among all affected family members extending from D16S451 to D1GS289, In order to extract maximum linkage information with missing individuals, incomplete informativeness with individual markers in this interval, and possible deviance from strict autosomal dominant inheritance, we performed nonparametric linkage analysis (NPL) and found a maximum NPL statistic corresponding to a P-value of .00098, close to the maximum conditional probability of linkage expected for a pedigree with this structure. Mutational analysis in this region specifically excludes expansion of the AT-rich minisatellite repeat FRA16B fragile site and the CAG trinucleotide repeat in the E2F-4 transcription factor. The ''repeat expansion detection'' method, capable of detecting dynamic mutation associated with anticipation, more generally excludes large CAG repeat expansion as a cause of leukemia in this family.

CFTR Polymorphisms in Patients with Alcoholic Chronic Pancreatitis

Introduction: Pancreas susceptibility to alcohol is variable and only 5-10% of chronic alcohol abusers develop chronic pancreatitis; the role of genetic factors in this process is unknown. The CFTR gene encodes a protein that acts on epithelial cells and plays a key role in normal exocrine pancreatic function. Methods: This study investigated the frequency of polymorphisms in intron 8 of the CFTR gene in patients with alcoholic chronic pancreatitis. Three groups of patients were studied: group A-68 adult alcoholics with a diagnosis of chronic pancreatitis; group B-68 adult alcoholics without pancreatic disease or liver cirrhosis and group C-104 healthy nonalcoholic adults. Results: T5/T7 genotype was more frequent in group A (11.8%) than in group B (2.9%) (p = 0.0481), and there was no statistical difference when groups A and C (5.8%) were compared (p = 0.1317). The haplotype combination (TG) 10-T7/(TG) 11-T7 was more frequent in groups B (23.5%) and C (20.2%) than in group A (7.3%) (p = 0.0080 and 0.0162). Conclusion: There are differences when these three groups are compared and individuals with T5/T7 genotype might have a greater risk of developing chronic pancreatitis when they become chronic alcoholics. Copyright (C) 2008 S. Karger AG, Basel and IAP

The GHEP-EMPOP collaboration on mtDNA population data-A new resource for forensic casework

Mitochondrial DNA (mtDNA) population data for forensic purposes are still scarce for some populations, which may limit the evaluation of forensic evidence especially when the rarity of a haplotype needs to be determined in a database search. In order to improve the collection of mtDNA lineages from the Iberian and South American subcontinents, we here report the results of a collaborative study involving nine laboratories from the Spanish and Portuguese Speaking Working Group of the International Society for Forensic Genetics (GHEP-ISFG) and EMPOP. The individual laboratories contributed population data that were generated throughout the past 10 years, but in the majority of cases have not been made available to the scientific community. A total of 1019 haplotypes from Iberia (Basque Country, 2 general Spanish populations, 2 North and 1 Central Portugal populations), and Latin America (3 populations from Sao Paulo) were collected, reviewed and harmonized according to defined EMPOP criteria. The majority of data ambiguities that were found during the reviewing process (41 in total) were transcription errors confirming that the documentation process is still the most error-prone stage in reporting mtDNA population data, especially when performed manually. This GHEP-EMPOP collaboration has significantly improved the quality of the individual mtDNA datasets and adds mtDNA population data as valuable resource to the EMPOP database (www.empop.org). (C) 2010 Elsevier Ireland Ltd. All rights reserved.

VKORC1 polymorphisms in Brazilians: comparison with the Portuguese and Portuguese-speaking Africans and pharmacogenetic implications

Aims: The heterogeneity of the Brazilian population renders the extrapolation of pharmacogenomic data derived from well-defined ethnic groups inappropriate. We investigated the influence of self-reported `race/color`, geographical origin and genetic ancestry on the distribution of four VKORC1 SNPs and haplotypes in Brazilians. Comparative data were obtained from two major ancestral roots of Brazilians: Portuguese and Africans from former Portuguese colonies. Materials & methods: A total of 1037 healthy adults Brazilians, recruited at four different geographical regions and self identified as white, brown or black (race/color categories), 89 Portuguese and 216 Africans from Angola and Mozambique were genotyped for the VKORC1 3673G>A (rs9923231), 5808T>G (rs2884737), 6853G>C (rs8050894) and 9041G>A (rs7294) polymorphisms using TaqMan (R) (Applied Biosystems, CA, USA) assays. VKORC1 haplotypes were statistically inferred using the haplo.stats software. We inferred the statistical association between the distribution of the VKORC1 polymorphisms among Brazilians and self-reported color, geographical region and genetic ancestry by fitting multinomial log linear models via neural networks. Individual proportions of European and African ancestry were used to assess the impact of genetic admixture on the frequency distribution of VKORC1 polymorphisms among Brazilians, and for the comparison of Brazilians with Portuguese and Africans. Results: The frequency distribution of the 3673G>A and 5808T>G polymorphisms, and VKORC1 haplotypes among Brazilians varies across geographical regions, within self-reported color categories and according to the individual proportions of European and African genetic ancestry. Notably, the frequency of the warfarin sensitive VKORC1 3673A allele and the distribution of VKORC1 haplotypes varied continuously as the individual proportion of European ancestry increased in the entire cohort, independently of race/color categorization and geographical origin. Brazilians with more than 80% African ancestry differ significantly from Angolans and Mozambicans in frequency of the 3673G>A, 5808T>G and 6853G>C polymorphisms and haplotype distribution, whereas no such differences are observed between Brazilians with more than 90% European ancestry and Portuguese individuals. Conclusion: The diversity of the Brazilian population, evident in the distribution of VKORC1 polymorphisms, must be taken into account in the design of pharmacogenetic clinical trials and dealt with as a continuous variable. Warfarin dosing algorithms that include `race` terms defined for other populations are clearly not applicable to the heterogeneous and extensively admixed Brazilian population.

Association Between Polymorphisms in GRIK2 Gene and Obsessive-Compulsive Disorder: A Family-Based Study

Several studies support a genetic influence on obsessive-compulsive disorder (OCD) etiology. The role of glutamate as an important neurotransmitter affecting OCD pathophysiology has been supported by neuroimaging, animal model, medication, and initial candidate gene studies. Genes involved in glutamatergic pathways, such as the glutamate receptor, ionotropic, kainate 2 (GRIK2), have been associated with OCD in previous studies. This study examines GRIK2 as a candidate gene for OCD susceptibility in a family-based approach. Probands had full DSM-IV diagnostic criteria for OCD. Forty-seven OCD probands and their parents were recruited from tertiary care OCD specialty clinics from France and USA. Genotypes of single nucleotide polymorphism (SNP) markers and related haplotypes were analyzed using Haploview and FBAT software. The polymorphism at rs1556995 (P = 0.0027; permuted P-value = 0.03) was significantly associated with the presence of OCD. Also, the two marker haplotype rs1556995/rs1417182, was significantly associated with OCD (P = 0.0019, permuted P-value = 0.01). This study supports previously reported findings of association between proximal GRIK2 SNPs and OCD in a comprehensive evaluation of the gene. Further study with independent samples and larger sample sizes is required.

Evaluation of RET polymorphisms in a six-generation family with G533C RET mutation: specific RET variants may modulate age at onset and clinical presentation

P>Context We previously described a six-generation family with G533C RET mutation and medullary thyroid carcinoma, in the largest family reported do date. Of particular interest, phenotype variability regarding the age of onset and clinical presentation of the disease, was observed. Objective We evaluate whether single SNPs within RET oncogene or haplotype comprising the RET variants (defined by Haploview) could predispose to early development of MTC in this family and influence the clinical manifestation. Design Eight SNPs were selected based on their previous association with the clinical course of hereditary or sporadic MTC, in particular promoting an early onset of disease. The variants were initially tested in 77 G533C-carriers and 100 controls using either PCR-direct sequencing or PCR-RFLP. Association between a SNP or haplotype and age at diagnosis or presence of lymph node metastasis was tested in 34 G533C-carries with MTC. Different bioinformatic tools were used to evaluate the potential effects on RNA splicing. Results An association was found between IVS1-126G > T and age at diagnosis. The variant [IVS8 +82A > G; 85-86 insC] was associated with the presence of lymph node metastases at diagnosis. In silico analysis suggested that this variant may induce abnormal splicing. This in silico analysis predicted that the [IVS8 +82A > G; 85-86 insC] could alter the splicing by disrupting and/or creating exonic splicing enhancer motifs. Conclusions We here identified two RET variants that were associated with phenotype variability in G533C-carriers, which highlights the fact that the modifier effect of a variant might depend on the type of mutation.

Role of promoter polymorphisms in the plasma glutathione peroxidase (GPx-3) gene as a risk factor for cerebral venous thrombosis

Background and Purpose-Plasma glutathione peroxidase (GPx-3) is a major antioxidant enzyme in plasma and the extracellular space that scavenges reactive oxygen species produced during normal metabolism or after oxidative insult. A deficiency of this enzyme increases extracellular oxidant stress, promotes platelet activation, and may promote oxidative posttranslational modification of fibrinogen. We recently identified a haplotype (H-2) in the GPx-3 gene promoter that increases the risk of arterial ischemic stroke among children and young adults. Methods-The aim of this study is to identify possible relationships between promoter haplotypes in the GPx-3 gene and cerebral venous thrombosis (CVT). We studied the GPx-3 gene promoter from 23 patients with CVT and 123 young controls (18 to 45 years) by single-stranded conformational polymorphism and sequencing analysis. Results-Over half of CVT patients (52.1%) were heterozygous (H1H2) or homozygous (H2H2) carriers of the H-2 haplotype compared with 12.2% of controls, yielding a more than 10-fold independent increase in the risk of CVT (OR=10.7; 95% CI, 2.70 to 42.36; P<0.0001). Among women, the interaction of the H2 haplotype with hormonal risk factors increased the OR of CVT to almost 70 (P<0.0001). Conclusions-These findings show that a novel GPx-3 promoter haplotype is a strong, independent risk factor for CVT. As we have previously shown that this haplotype is associated with a reduction in transcriptional activity, which compromises antioxidant activity and antithrombotic benefits of the enzyme, these results suggest that a deficiency of GPx-3 leads to a cerebral venous thrombophilic state.

Variants in the promoter region of IKBL/NFKBIL1 gene may mark susceptibility to the development of chronic Chagas` cardiomyopathy among Trypanosoma cruzi-infected individuals

Chagas` disease, caused by Trypanosoma cruzi, is an inflammatory disorder leading to chronic Chagas cardiomyopathy (CCC). Only one third of T cruzi-infected individuals progress to CCC while the others are considered asymptomatic (ASY). The human inhibitory kappa B-like gene (KBLINFKBIL1), homologous to the I kappa B family of proteins that regulate the NF kappa B family of transcription factors, is suggested as a putative inhibitor of NFKB. We investigated two functional polymorphisms, -62A/T and -262A/G, in the promoter of IKBL by PCR-RFLP analysis in 169 patients with CCC and 76 ASY. Genotype distributions for both -62A/T and -262A/G differed between the CCC and ASY (X-2 = 7.3; P = 0.025 and X-2 = 6.8; P = 0.03, respectively). Subjects, homozygous for the -62A allele, had three-fold risk of developing CCC compared with those carrying the TT genotype (P = 0.0095; Odds Ratio [OR] = 2.9; [95% CI 1.2-7.3]). Similar trend was observed for the -262A homozygotes (P = 0.005; OR = 2.7 [95% CI 1.3-6.0]. The haplotype -262A -62A was prevalent in patients with CCC (40% versus 24%; OR 2.1 [95% C1 1.4-3.3j; Pc = 0.00 14). The I kappa BL locus itself or another critical gene in this region may confer susceptibility to the development of CCC. (C) 2007 Elsevier Ltd. All rights reserved.

In silico analysis of microRNAS targeting the HLA-G 3 ` untranslated region alleles and haplotypes

It has been reported that microRNAs (miRNA) may have allele-specific targeting for the 3` untranslated region (3` UTR) of the HLA-G locus. In a previous study, we reported 11 3`UTR haplotypes encompassing the 14-bp insertion/deletion polymorphism and seven SNPs (+3003 T/C, +3010 C/G, +3027 C/A, +3035 C/T, +3142 C/G, +3187A/G,and +3196 C/G), of which only the +3142 C/G SNP has been reported to influence the binding of miRNAs. Using bioinformatics analyses, we identified putative miRNA-binding sites considering the haplotypes encompassing these eight polymorphic sites, and we ranked the lowest free energies that could potentially lead to an mRNA degradation or translational repression. When a specific haplotype or a particular SNP was associated with a miRNA-binding site, we defined a free energy difference of 4 kcal/mol between alleles to classify them energetically distant. The best results were obtained for the miR-513a-5p, miR-518c*, miR-1262 and miR-92a-1*, miR-92a-2*, miR-661, miR-1224-5p, and miR-433 miRNAs, all influencing one or more of the +3003, +3010, +3027, and +3035 SNPs. The miR-2110, miR-93, miR-508-5p, miR-331-5p, miR-616, miR-513b, and miR-589* miRNAs targeted the 14-bp fragment region, and miR-148a, miR-19a*, miR-152, mir-148b,and miR-218-2 also influenced the +3142C/G polymorphism. These results suggest that these miRNAs might play a relevant role on the HLA-G expression pattern. (C) 2009 Published by Elsevier Inc. on behalf of American Society for Histocompatibility and Immunogenetics.

HLA-G polymorphisms in women with squamous intraepithelial lesions harboring human papillomavirus

Human papillomavirus (HPV) infection is etiologically associated with low-(LSIL) and high-grade squamous intraepithelial lesions (HSIL) and with cervical cancer. The progression or regression of the lesions may depend, among other factors, on the host heritable immune response. Because human leukocyte antigen (HLA)-G molecules are involved in the modulation of innate and adaptive immune responses, and because no previous studies have evaluated HLA-G polymorphism in patients with SIL, we conducted a study to assess the association between HLA-G polymorphisms and cervical lesions harboring HPV infection. Cervico-vaginal scrapings and blood samples were collected from 125 women with SIL (68 LSIL and 57 HSIL) and from 94 healthy women without HPV infection and cytological abnormalities. HPV type and HLA-G polymorphisms in exons 2, 3 and 8 (14 bp insertion/deletion) were evaluated by PCR methodology, and digested with restriction endonucleases. The Genepop software and the EM and PHASE algorithms were used for statistical analysis. A significant protective association was observed between the presence of the G*0103 allele and SIL and between the G0101/G0104 genotype and HSIL in the group of patients compared to control. The presence of the G0104/+14 bp and G0104/-14 bp haplotypes conferred susceptibility to SIL compared to control. In addition, patients possessing the G0104/+14 bp haplotype and harboring HPV-16 and -18 co-infections were particularly associated with HSIL. These findings suggest that HLA-G polymorphisms may be associated with HPV infection and SIL, consequently representing a profile of predisposition to cervical cancer. Modern Pathology (2009) 22, 1075-1082; doi: 10.1038/modpathol.2009.67; published online 1 May 2009

HLA-G polymorphism and transitional cell carcinoma of the bladder in a Brazilian population

The morphologic appearance and clinical behavior of the human urinary bladder papillary transitional cell carcinoma (TCC) probably result from a complex interaction between carcinogenic insults and host resistance during the patient`s life. While the main recognized risk factors are of environmental origin (e.g. smoking), relatively little information exists about the susceptibility to TCC development. The human leukocyte antigen G (HLA-G) molecule plays an important role in immune response regulation and has been implicated in the inhibition of the cytolytic function of natural killer and cytotoxic T cells. Several lines of evidence indicate that HLA-G polymorphisms influence the expression level and production of different HLA-G isoforms. The aim of this study was to explore a possible influence of the HLA-G polymorphism on the susceptibility to urinary bladder TCC development and progression in smokers and nonsmokers Brazilian subjects. The HLA-G locus was found to be associated with susceptibility to TCC development and progression. The G*0104 allelic group (specially the G*010404 allele) and the G*0103 allele were associated with a tobacco-dependent influence on TCC development. The G*0104 group was associated with progression to high-grade tumors, irrespective of smoking habit, while the G*0103 allele was associated to high-grade tumor only in smoking patients. Our results are an evidence that the HLA-G locus itself, or as part of an extended haplotype encompassing this chromosome region (particularly the HLA-A given the high linkage disequilibrium observed between them in this data series), may be associated with TCC susceptibility and tumor progression, suggesting a tobacco-dependent influence of these polymorphisms.