Transmission test for linkage disequilibrium: the insulin gene region and insulin-dependent diabetes mellitus (IDDM).

A population association has consistently been observed between insulin-dependent diabetes mellitus (IDDM) and the "class 1" alleles of the region of tandem-repeat DNA (5'' flanking polymorphism [5''FP]) adjacent to the insulin gene on chromosome 11p. This finding suggests that the insulin gene region contains a gene or genes contributing to IDDM susceptibility. However, several studies that have sought to show linkage with IDDM by testing for cosegregation in affected sib pairs have failed to find evidence for linkage. As means for identifying genes for complex diseases, both the association and the affected-sib-pairs approaches have limitations. It is well known that population association between a disease and a genetic marker can arise as an artifact of population structure, even in the absence of linkage. On the other hand, linkage studies with modest numbers of affected sib pairs may fail to detect linkage, especially if there is linkage heterogeneity. We consider an alternative method to test for linkage with a genetic marker when population association has been found. Using data from families with at least one affected child, we evaluate the transmission of the associated marker allele from a heterozygous parent to an affected offspring. This approach has been used by several investigators, but the statistical properties of the method as a test for linkage have not been investigated. In the present paper we describe the statistical basis for this "transmission test for linkage disequilibrium" (transmission/disequilibrium test [TDT]). We then show the relationship of this test to tests of cosegregation that are based on the proportion of haplotypes or genes identical by descent in affected sibs. The TDT provides strong evidence for linkage between the 5''FP and susceptibility to IDDM. The conclusions from this analysis apply in general to the study of disease associations, where genetic markers are usually closely linked to candidate genes. When a disease is found to be associated with such a marker, the TDT may detect linkage even when haplotype-sharing tests do not.     

Multifactorial diseases: a nightmare for the geneticist

Common diseases are often familial, but they do not show in most families, a simple pattern of inheritance. In a few families these diseases may be caused by a mutation in a single gene. In most families these diseases are multifactorial, they result from a complex interaction between a genetic component which is often polygenic and many environmental factors. Two major, model free, methods are used to locate and identify susceptibility genes that predispose to multifactorial diseases. The first is a non parametric linkage analysis that relies on affected sib pairs, or an affected pedigree member, the second method is association studies which looks for increase frequency of particular alleles or genotypes in affected compared with unaffected individuals in the population. Most of the results have not been replicated, identifying susceptibility genes is proving much more difficult than most geneticists imagined 20 years ago. The main reason for this irreproducibility is genetic heterogeneity.     

高血压易感基因的分子进化

利用家系连锁分析、候选基因法及全基因组关联研究均未能有效发现普通人群的高血压易感基因或位点。遗传学研究表明, 人类许多疾病易感性的形成与走出非洲时的环境适应性进化密切相关, 这为高血压遗传学研究提供了新思路。文章系统综述了高血压易感基因分子进化研究的理论基础和最新进展, 介绍了本研究小组运用分子进化思路在中国汉族人群高血压遗传学研究中的发现, 对未来的研究方向进行了展望, 以期为高血压和其他疾病的遗传学研究提供参考。     

Genetics of LDL particle heterogeneity: from genetic epidemiology to DNA-based variations

Substantial evidence exists suggesting that small, dense LDL particles are associated with an increased risk of coronary heart disease. This disease-related risk factor is recognized to be under both genetic and environmental influences. Several studies have been conducted to elucidate the genetic architecture underlying this trait, and a review of this literature seems timely. The methods and strategies used to determine its genetic component and to identify the genes have greatly changed throughout the years owing to the progress made in genetic epidemiology and the influence of the Human Genome Project. Heritability studies, complex segregation analyses, candidate gene linkage and association studies, genome-wide linkage scans, and animal models are all part of the arsenal to determine the susceptibility genes. The compilation of these studies clearly revealed the complex genetic nature of LDL particles. This work is an attempt to summarize the growing evidence of genetic control on LDL particle heterogeneity with the aim of providing a concise overview in one read.     

Molecular genetic approaches for studying the etiology of diabetic nephropathy

A critical challenge faced by clinical nephrologists today is the escalating number of patients developing end stage renal disease, a major proportion of which is attributed to diabetic nephropathy (DN). The need for new measures to prevent and treat this disease cannot be overemphasized. To this end, modern genetic approaches provide powerful tools to investigate the etiology of DN. Human studies have already established the importance of genetic susceptibility for DN. Several major susceptibility loci have been identified using linkage studies. In addition, linkage studies in rodents have pinpointed promising chromosomal segments that influence renal traits. Besides augmenting our understanding of disease pathogenesis, these animal studies may facilitate the cloning of disease susceptibility genes in man through the identification of homologous regions that contribute to renal disease. In human diabetes, various genes have been evaluated for their risk contribution to DN. This widespread strategy has been propelled by our knowledge of the glucose-activated pathways underlying DN. Evidence has emerged that a true association does indeed exist for some candidate genes. Furthermore, the in vivo manipulation of gene expression has shown that these genes can modify features of DN in transgenic and knockout rodent models, thus corroborating the findings from human association studies. Still, the exact molecular mechanisms involving these genes remain to be fully elucidated. This formidable task may be accomplished by continuing to harness the synergy between human and experimental genetic approaches. In this respect, our review provides a first synthesis of the current literature to facilitate this challenging effort.     

Normative genetic profiles of RAAS pathway gene polymorphisms in North Indian and South Indian populations

Population-based genetic association studies, popularly known as case-control studies, have continued to be the most preferred method for deciphering the genetic basis of various complex diseases, even in the post-human genome sequencing era. However, interpopulation differences in allele, genotype, and haplotype frequencies and linkage disequilibrium patterns lead to inconsistent results in candidate gene association studies. Therefore, for any meaningful disease association study, knowledge of the normative genetic background of the baseline population is a prerequisite. In addition, such genetic variation data also provide a ready-made menu of allele frequencies and linkage disequilibrium patterns of various polymorphisms in specific candidate genes in a particular population, which is a useful reference for further genetic association studies. Such genetic variation data are lacking for the Indian population, which represents about one-sixth of the world's population. In the present study we have reported the allele, genotype, and haplotype frequencies, Hardy-Weinberg equilibrium status, and linkage disequilibrium patterns of 12 polymorphisms in six candidate genes from the renin-angiotensin-aldosterone system among Indians. Because of their different history of origin, the Indian population is broadly divided into two subpopulations: North Indians (Caucasian Europeans) and South Indians (Dravidians). Considering this well-documented difference in gene pools, we have presented a comparative account of the normative genetic data of North Indian and South Indian populations with at least four individuals of urban and suburban origin from each of the representative states of northern and southern India.     

The transmission/disequilibrium test: history, subdivision, and admixture.

Disease association with a genetic marker is often taken as a preliminary indication of linkage with disease susceptibility. However, population subdivision and admixture may lead to disease association even in the absence of linkage. In a previous paper, we described a test for linkage (and linkage disequilibrium) between a genetic marker and disease susceptibility; linkage is detected by this test only if association is also present. This transmission/disequilibrium test (TDT) is carried out with data on transmission of marker alleles from parents heterozygous for the marker to affected offspring. The TDT is a valid test for linkage and association, even when the association is caused by population subdivision and admixture. In the previous paper, we did not explicitly consider the effect of recent history on population structure. Here we extend the previous results by examining in detail the effects of subdivision and admixture, viewed as processes in population history. We describe two models for these processes. For both models, we analyze the properties of (a) the TDT as a test for linkage (and association) between marker and disease and (b) the conventional contingency statistic used with family data to test for population association. We show that the contingency test statistic does not have a chi 2 distribution if subdivision or admixture is present. In contrast, the TDT remains a valid chi 2 statistic for the linkage hypothesis, regardless of population history.     

Multiple genes for essential-hypertension susceptibility on chromosome 1q

Essential hypertension, defined as elevated levels of blood pressure (BP) without any obvious cause, is a major risk factor for coronary heart disease, stroke, and renal disease. BP levels and susceptibility to development of essential hypertension are partially determined by genetic factors that are poorly understood. Similar to other efforts to understand complex, non-Mendelian phenotypes, genetic dissection of hypertension-related traits employs genomewide linkage analyses of families and association studies of patient cohorts, to uncover rare and common disease alleles, respectively. Family-based mapping studies of elevated BP cover the large intermediate ground for identification of genes with common variants of significant effect. Our genomewide linkage and candidate-gene-based association studies demonstrate that a replicated linkage peak for BP regulation on human chromosome 1q, homologous to mouse and rat quantitative trait loci for BP, contains at least three genes associated with BP levels in multiple samples: ATP1B1, RGS5, and SELE. Individual variants in these three genes account for 2-5-mm Hg differences in mean systolic BP levels, and the cumulative effect reaches 8-10 mm Hg. Because the associated alleles in these genes are relatively common (frequency >5%), these three genes are important contributors to elevated BP in the population at large.     

Genetic variation at the matrix metalloproteinase-9 locus on chromosome 20q12.2–13.1

Allelic association methods are better suited than linkage analysis for mapping of susceptibility genes that confer modest increases in risk in complex diseases. In both family- and population-based association studies, it is very useful to have prior knowledge of all sequence variants and the degree of linkage disequilibrium in a candidate gene region. In this study, we scanned sequence variants in a 2.2-kb promoter sequence and all 13 exons (totalling 3.3 kb) of the matrix metalloproteinase-9 gene, which is associated with coronary heart disease and a candidate for other diseases involving connective tissue remodelling, such as cancer metastasis. The sequences had a total of ten variable sites, four in the promoter, five in the coding region (three of which alter the amino acid encoded) and one in the 3' untranslated sequence. Sequence inspection suggests that some of the variants will have a functional impact on either level of expression or enzymatic activity. Tight linkage disequilibrium was detected between variants across the entire length of the gene (approximately 9 kb), and frequencies of different haplotypes were determined. The data provide an essential tool for studies of the possible contribution of genetic variation at the matrix metalloproteinase-9 locus to genetically determined susceptibility to a number of important diseases. The results also provide experimental data on the extent of linkage disequilibrium in the general population, which is yet to be resolved.     

基因水平的关联分析方法

全基因组关联研究(Genome wide association study, GWAS)已经在国内外的医学遗传学研究中得到广泛应用, 但是GWAS数据中所蕴含的与多基因复杂性状疾病机制相关的丰富信息尚未得到深度挖掘。近年来, 研究者采用生物网络分析和生物通路分析等生物信息学和生物统计学手段分析GWAS数据, 并探索潜在的疾病机制。生物网络分析和生物通路分析主要是以基因为单位进行的, 因此必须在分析前将基因上全部或者部分单个单核苷酸多态性(Single nucleotide polymorphism, SNP)的遗传关联结果综合起来, 即基因水平的关联分析。基因水平的关联分析需要考虑单个SNP的遗传关联、基因上SNP数量和SNP之间的连锁不平衡结构等多种因素, 因此不仅在遗传学的概念上也在统计方法方面具有一定的复杂性和挑战性。文章对基因水平的关联分析的研究进展、原理和应用进行了综述。     

Immunogenetics of leishmanial and mycobacterial infections: the Belem Family Study.

In the 1970s and 1980s, analysis of recombinant inbred, congenic and recombinant haplotype mouse strains permitted us to effectively ''scan'' the murine genome for genes controlling resistance and susceptibility to leishmanial infections. Five major regions of the genome were implicated in the control of infections caused by different Leishmania species which, because they show conserved synteny with regions of the human genome, immediately provides candidate gene regions for human disease susceptibility genes. A common intramacrophage niche for leishmanial and mycobacterial pathogens, and a similar spectrum of immune response and disease phenotypes, also led to the prediction that the same genes/candidate gene regions might be responsible for genetic susceptibility to mycobacterial infections such as leprosy and tuberculosis. Indeed, one of the murine genes (Nramp1) was identified for its role in controlling a range of intramacrophage pathogens including leishmania, salmonella and mycobacterium infections. In recent studies, multicase family data on visceral leishmaniasis and the mycobacterial diseases, tuberculosis and leprosy, have been collected from north-eastern Brazil and analysed to determine the role of these candidate genes/regions in determining disease susceptibility. Complex segregation analysis provides evidence for one or two major genes controlling susceptibility to tuberculosis in this population. Family-based linkage analyses (combined segregation and linkage analysis; sib-pair analysis), which have the power to detect linkage between marker loci in candidate gene regions and the putative disease susceptibility genes over 10-20 centimorgans, and transmission disequilibrium testing, which detects allelic associations over 1 centimorgan (ca. 1 megabase), have been used to examine the role of four regions in determining disease susceptibility and/or immune response phenotype. Our results demonstrate: (i) the major histocompatibility complex (MHC: H-2 in mouse, HLA in man: mouse chromosome 17/human 6p; candidates class II and class III including TNF alpha/beta genes) shows both linkage to, and allelic association with, leprosy per se, but is only weakly associated with visceral leishmaniasis and shows neither linkage to nor allelic association with tuberculosis; (ii) no evidence for linkage between NRAMP1, the positionally cloned candidate for the murine macrophage resistance gene Ity/Lsh/Bcg (mouse chromosome 1/human 2q35), and susceptibility to tuberculosis or visceral leishmaniasis could be demonstrated in this Brazilian population; (iii) the region of human chromosome 17q (candidates NOS2A, SCYA2-5) homologous with distal mouse chromosome 11, originally identified as carrying the Scl1 gene controlling healing versus nonhealing responses to Leishmania major, is linked to tuberculosis susceptibility; and (iv) the ''T helper 2'' cytokine gene cluster (proximal murine chromosome 11/human 5q; candidates IL4, IL5, IL9, IRF1, CD14) controlling later phases of murine L. major infection, is not linked to human disease susceptibility for any of the three infections, but shows linkage to and highly significant allelic association with ability to mount an immune response to mycobacterial antigens. These studies demonstrate that the ''mouse-to-man'' strategy, refined by our knowledge of the human immune response to infection, can lead to the identification of important candidate gene regions in man.     

Introduction to Deep Sequencing and Its Application to Drug Addiction Research with a Focus on Rare Variants

Through linkage analysis, candidate gene approach, and genome-wide association studies (GWAS), many genetic susceptibility factors for substance dependence have been discovered such as the alcohol dehydrogenase gene (ALDH2) for alcohol dependence (AD) and nicotinic acetylcholine receptor (nAChR) subunit variants on chromosomes 8 and 15 for nicotine dependence (ND). However, these confirmed genetic factors contribute only a small portion of the heritability responsible for each addiction. Among many potential factors, rare variants in those identified and unidentified susceptibility genes are supposed to contribute greatly to the missing heritability. Several studies focusing on rare variants have been conducted by taking advantage of next-generation sequencing technologies, which revealed that some rare variants of nAChR subunits are associated with ND in both genetic and functional studies. However, these studies investigated variants for only a small number of genes and need to be expanded to broad regions/genes in a larger population. This review presents an update on recently developed methods for rare-variant identification and association analysis and on studies focused on rare-variant discovery and function related to addictions.     

不宁腿综合征遗传学研究进展

不宁腿综合征(Restless legs syndrome, RLS)遗传学研究近年来获得了许多重要的进展, 极大地丰富了对于这种疾病分子机制的认识。RLS是一种常见的复杂疾病, 几个遗传流行病学和双生子研究对RLS遗传组分进行了剖析, 说明RLS是一个遗传性很强的性状, 其遗传力约为50%。采用基于模型的连锁分析方法或者是不依赖于模型的连锁分析方法目前已定位了5个重要的RLS疾病连锁位点: 12q13-23, 14q13-21, 9p24-22, 2q33和20p13, 为定位克隆RLS致病基因或者易感基因提供了连锁图谱。最新基于高通量的SNPs分型平台开展的全基因组分析确立3个与RLS显著关联的区域: 6p21.2, 2p14和15q23。文章结合作者近年来从事不宁腿综合征遗传学的研究工作, 对该领域的重要成果进行了汇总和评述。     

Network-Based Multiple Sclerosis Pathway Analysis with GWAS Data from 15,000 Cases and 30,000 Controls

Multiple sclerosis (MS) is an inflammatory CNS disease with a substantial genetic component, originally mapped to only the human leukocyte antigen (HLA) region. In the last 5 years, a total of seven genome-wide association studies and one meta-analysis successfully identified 57 non-HLA susceptibility loci. Here, we merged nominal statistical evidence of association and physical evidence of interaction to conduct a protein-interaction-network-based pathway analysis (PINBPA) on two large genetic MS studies comprising a total of 15,317 cases and 29,529 controls. The distribution of nominally significant loci at the gene level matched the patterns of extended linkage disequilibrium in regions of interest. We found that products of genome-wide significantly associated genes are more likely to interact physically and belong to the same or related pathways. We next searched for subnetworks (modules) of genes (and their encoded proteins) enriched with nominally associated loci within each study and identified those modules in common between the two studies. We demonstrate that these modules are more likely to contain genes with bona fide susceptibility variants and, in addition, identify several high-confidence candidates (including BCL10, CD48, REL, TRAF3, and TEC). PINBPA is a powerful approach to gaining further insights into the biology of associated genes and to prioritizing candidates for subsequent genetic studies of complex traits.     

HLA-linked rheumatoid arthritis.

Twenty-eight pedigrees were ascertained through pairs of first-degree relatives diagnosed with rheumatoid arthritis (RA). RA was confirmed in 77 pedigree members including probands; the absence of disease was verified in an additional 261 pedigree members. Pedigree members were serologically typed for HLA. We used likelihood analysis to statistically characterize the HLA-linked RA susceptibility locus. The genetic model assumed tight linkage to HLA. The analysis supported the existence of an HLA-linked RA susceptibility locus, estimated the susceptibility allele frequency as 2.16%, and estimated the lifetime penetrance as 41% in male homozygotes and as 48% in female homozygotes. Inheritance was recessive in males and was nearly recessive in females. In addition, the analysis attributed 78% of the variance within genotypes to genetic or environmental effects shared by siblings. The genetic model inferred in this analysis is consistent with previous association, linkage, and familial aggregation studies of RA. The inferred HLA-linked RA susceptibility locus accounts for approximately one-half of familial RA, although it accounts for only approximately one-fifth of the RA in the population. Although other genes may account for the remaining familial RA, a large portion of RA cases may occur sporadically.     

No genetic association of the human prolyl endopeptidase gene in the Dutch celiac disease population

Celiac disease (CD) is a complex genetic disorder of the small intestine. The DQ2/DQ8 human leucocyte antigen (HLA) genes explain approximately 40% of the genetic component of the disease, but the remaining non-HLA genes have not yet been identified. The key environmental factor known to be involved in the disease is gluten, a major protein present in wheat, barley, and rye. Integrating microarray data and linkage data from chromosome 6q21-22 revealed the prolyl endopeptidase (PREP) gene as a potential CD candidate in the Dutch population. Interestingly, this gene encodes for the only enzyme that is able to cleave the proline-rich gluten peptides. To investigate the role of the human PREP gene as a primary genetic factor in CD, we conducted gene expression, sequence analysis, and genetic association studies of the PREP gene and determined PREP enzyme activity in biopsies from CD patients and controls. Sequence analysis of the coding region of the PREP gene revealed two novel polymorphisms. Genetic association studies using two novel polymorphisms and three known PREP variants excluded a genetic association between PREP and CD. Determination of PREP activity revealed weak but significant differences between treated and untreated CD biopsies (P < 0.05). Our results from the association study indicate that PREP is not a causative gene for CD in the Dutch population. These are further supported by the activity determinations in which we observed no differences in PREP activity between CD patients and controls.     

Vitiligo: pathomechanisms and genetic polymorphism of susceptible genes

Vitiligo is a depigmenting disorder resulting from the loss of melanocytes in the skin and affects 1-4% of the world population. Incidence of vitiligo is found to be 0.5-2.5% in India with a high prevalence of 8.8% in Gujarat and Rajasthan states. The cellular and molecular mechanisms that lead to melanocyte destruction in this disorder are not yet been fully elucidated. Genetic factors, neural factors, toxic ROS metabolites, autoantibodies and autoreactive T lymphocytes may be the causative agents for the selective destruction of melanocytes. Three major hypotheses of pathogenesis of vitiligo are neural, autoimmune and oxidative stress hypotheses, however none of them explains the pathogenesis of vitiligo in toto. Genetics of vitiligo is characterized by incomplete penetrance, multiple susceptibility loci and genetic heterogeneity. Recent advances in this field are linkage and association of candidate gene studies. The linkage and association studies provide a strong evidence for the presence of multiple vitiligo susceptibility genes on different chromosomes. Several candidate genes for vitiligo are identified from different populations. In this review, we have provide an overview of different hypotheses of vitiligo pathogenesis, and discuss the recent advances in this field with special reference to linkage, association and candidate gene approach.     

Recent advances in the genetic investigation of osteoarthritis

Osteoarthritis (OA) demonstrates considerable clinical heterogeneity, generating heated debate over whether OA is a single disease or a complex mix of disparate diseases and concerning which tissues are principally involved in disease initiation and progression. Epidemiological studies have demonstrated a major genetic component to OA risk. However, these studies have also revealed differences in risk between males and females and for disease at different skeletal sites. This observation has resulted in the concept of genes for specific sites rather than a generalised OA phenotype. Recent breakthroughs have shed considerable light on the nature of OA genetic susceptibility. Many candidate genes have been confirmed, such as the interleukin-1 gene cluster and the oestrogen alpha-receptor gene ESR1. Genome-wide linkage scans have revealed several regions harbouring novel loci, some of which are beginning to yield their genes.     

Score tests of genetic association in the presence of linkage based on the additive genetic gamma frailty model

Nuclear families with multiple affected sibs are often collected for genetic linkage analysis of complex diseases. Once linkage evidence is established, dense markers are often typed in the linked region for genetic association analysis based on linkage disequilibrium (LD). Detection of association in the presence of linkage localizes disease genes more accurately than the methods that rely on linkage alone. However, test of association due to LD in the linked region needs to account for dependency of the allele transmissions to different sibs within a family. In this paper, we define a joint model for genetic linkage and association and derive the corresponding joint survival function of age of onset for the sibs within a sibship. The joint survival function is a function of both the inheritance vector and the genotypes at the candidate marker locus. Based on this joint survival function, we derive score tests for genetic association. The proposed methods utilize the phenotype data of all the sibs and have the advantages of family-based designs which can avoid the potential spurious association caused by population admixture. In addition, the methods can account for variable age of onset or age at censoring and possible covariate effects, and therefore provide important tools for modelling disease heterogeneity. Simulation studies and application to the data sets from the 12th Genetic Analysis Workshop indicate that the proposed methods have correct type 1 error rates and increased power over other existing methods for testing allelic association.