Genetic variability and linkage disequilibrium within the HLA-DP region: analysis of 15 different populations

In order to understand the forces governing the evolution of the genetic diversity in the HLA-DP molecule, polymerase chain reaction (PCR)-based methods were used to characterize genetic variation at the DPA1 and DPB1 loci encoding this heterodimer on 2,807 chromosomes from 15 different populations including individuals of African, Asian, Amerindian, Indian and European origin. These ethnically diverse samples represent a variety of population substructures and include small, isolated populations as well as larger, presumably admixed populations. Ten DPA1 and 39 DPB1 alleles were identified and observed on 87 distinct DP haplotypes, 34 of which were found to be in significant positive linkage disequilibrium in at least one population. Some haplotypes were found in all ethnic groups while others were confined to a single ethnic group or population. Strong positive global linkage disequilibrium (Wn) between DPA1 and DPB1 was present in all 15 populations. The African populations displayed the lowest values of Wn whereas the Amerindian populations displayed near absolute disequilibrium. Analysis of the distribution of haplotypes using the normalized deviate of the Ewens-Watterson homozygosity statistic, F, suggests that DP haplotypes encoding the functional heterodimer are subject to much lower degrees of balancing selection than other loci within the HLA region. Finally, neighbor joining tree analyses demonstrate the power of haplotype diversity for inferring the relationships between the different populations.     

Allelic association with SNPs: metrics, populations, and the linkage disequilibrium map

Comparison of different metrics, using three large samples of haplotypes from different populations, demonstrates that rho is the most efficient measure of association between pairs of single nucleotide polymorphisms (SNPs). Pairwise data can be modeled, using composite likelihood, to describe the decline in linkage disequilibrium with distance (the Malecot model). The evidence from more isolated populations (Finland, Sardinia) suggests that linkage disequilibrium extends to 427-893 kb but, even in samples representative of large heterogeneous populations, such as CEPH, the extent is 385 kb or greater. This suggests that isolated populations are not essential for linkage disequilibrium mapping of common diseases with SNPs. The in parameter of the Malecot model (recombination and time), evaluated at each SNP, indicates regions of the genome with extensive and less extensive disequilibrium (low and high values of in respectively). When plotted against the physical map, the regions with extensive and less extensive linkage disequilibrium may correspond to recombination cold and hot spots. This is discussed in relation to the Xq25 cytogenetic band and the HFE gene region.     

Sequence features in regions of weak and strong linkage disequilibrium

We use genotype data generated by the International HapMap Project to dissect the relationship between sequence features and the degree of linkage disequilibrium in the genome. We show that variation in linkage disequilibrium is broadly similar across populations and examine sequence landscape in regions of strong and weak disequilibrium. Linkage disequilibrium is generally low within approximately 15 Mb of the telomeres of each chromosome and noticeably elevated in large, duplicated regions of the genome as well as within approximately 5 Mb of centromeres and other heterochromatic regions. At a broad scale (100-1000 kb resolution), our results show that regions of strong linkage disequilibrium are typically GC poor and have reduced polymorphism. In addition, these regions are enriched for LINE repeats, but have fewer SINE, DNA, and simple repeats than the rest of the genome. At a fine scale, we examine the sequence composition of "hotspots" for the rapid breakdown of linkage disequilibrium and show that they are enriched in SINEs, in simple repeats, and in sequences that are conserved between species. Regions of high and low linkage disequilibrium (the top and bottom quartiles of the genome) have a higher density of genes and coding bases than the rest of the genome. Closer examination of the data shows that whereas some types of genes (including genes involved in immune response and sensory perception) are typically located in regions of low linkage disequilibrium, other genes (including those involved in DNA and RNA metabolism, response to DNA damage, and the cell cycle) are preferentially located in regions of strong linkage disequilibrium. Our results provide a detailed analysis of the relationship between sequence features and linkage disequilibrium and suggest an evolutionary justification for the heterogeneity in linkage disequilibrium in the genome.     

Genetic susceptibility factors in type 1 diabetes: linkage, disequilibrium and functional analyses

Continuing progress has been made in elucidating the genetic factors involved in type 1 diabetes (insulin-dependent diabetes mellitus [IDDM]) in the past year. Two genome scans suggested additional susceptibility intervals and provided supporting evidence for several previously reported linkages. Other studies focused on the confirmation of linkage using multipoint sibpair analyses with densely spaced markers and multiethnic collections of families. Although significant and consistent linkage evidence was reported for the susceptibility intervals IDDM8 (on human chromosome 6q27), IDDM4 (on 11q) and IDDM5 (on 6q25), evidence for most other intervals varies in different data sets — probably due to a weak effect of the disease genes, genetic heterogeneity or random variation. Linkage disequilibrium mapping has become an increasingly important tool for both the confirmation and fine-mapping of susceptibility intervals, as well as identification of etiological mutations. Functional studies indicate, firstly, that the susceptible and protective HLA class II molecules HLA-DR and -DQ bind and present nonoverlapping peptides and, secondly, that the variable number of tandem repeats at the 5′ end of the insulin gene (susceptibility interval IDDM2) regulates insulin expression in the thymus.     

A fine-scale comparison of the human and chimpanzee genomes: linkage, linkage disequilibrium and sequence analysis

We have performed a fine-scale comparative study of the human and chimpanzee genomes, using linkage, linkage disequilibrium and sequence analyses on microsatellite loci spanning a region of approximately 30 cM on human chromosome 4p. Our results extend the findings of previous studies that indicated virtually complete conservation between the human and chimpanzee genomes at the chromosomal and sub-chromosomal level and support the hypothesis, derived from previous analyses of mitochondrial DNA, that chimpanzee populations are more diverse than human ones. By sequencing several human and chimpanzee alleles of two microsatellites we showed that base substitutions that diminish the length of perfect repeats (but do not change allele sizes) are probably responsible for the low heterozygosity of these loci in chimpanzees; our results suggest that the evolutionary history of microsatellites should not be inferred from comparisons of mean allele lengths between populations or species.      

Comparative study of the haplotype structure and linkage disequilibrium of chromosome 1p36.2 region in the Korean and Japanese populations

The patterns of linkage disequilibrium (LD) in the human genome provide important information for disease gene mapping. LDs may vary depending on chromosomal regions and populations. We have compared LD and haplotypes defined by SNPs in the chromosome 1p36.2 region of the Korean and Japanese populations. Fifty-eight SNPs in about 418 kb ranging from tumor necrosis factor receptor 2 (TNFR2:TNFRSF1B) to procollagen-lysine, 2-oxoglutarate 5-dioxygenase (PLOD) gene were examined in 96 healthy Koreans and Japanese each by direct sequencing and fluorescence correlation spectroscopy combined with the PCR-sequence specific primer method (PCR-SSP-FCS), respectively. Upon pair-wise LD analysis, a total of 25 and 16 out of 58 SNPs greater than MAF 10% were included in LD blocks, encompassing almost 81 kb and 55 kb in total, in Koreans and Japanese, respectively. Both similarities and differences were observed in LD strength and haplotype frequencies between the populations. Considerable similarities were observed in the telomeric region where a long-range block of approximately 80 kb including three genes was found to have strong LDs in both Koreans and Japanese. Significant difference in LD strength was present near the TNFR2 region between the Japanese and Korean populations.     

The linkage disequilibrium maps of three human chromosomes across four populations reflect their demographic history and a common underlying recombination pattern

The extent and patterns of linkage disequilibrium (LD) determine the feasibility of association studies to map genes that underlie complex traits. Here we present a comparison of the patterns of LD across four major human populations (African-American, Caucasian, Chinese, and Japanese) with a high-resolution single-nucleotide polymorphism (SNP) map covering almost the entire length of chromosomes 6, 21, and 22. We constructed metric LD maps formulated such that the units measure the extent of useful LD for association mapping. LD reaches almost twice as far in chromosome 6 as in chromosomes 21 or 22, in agreement with their differences in recombination rates. By all measures used, out-of-Africa populations showed over a third more LD than African-Americans, highlighting the role of the population's demography in shaping the patterns of LD. Despite those differences, the long-range contour of the LD maps is remarkably similar across the four populations, presumably reflecting common localization of recombination hot spots. Our results have practical implications for the rational design and selection of SNPs for disease association studies.     

Diversity of MICA and linkage disequilibrium with HLA-B in two North American populations

The MICA gene has a high degree of polymorphism. Allelic variation of MICA may influence binding of these ligands to the NK cell receptor NKG2D and may affect organ transplantation and/or disease pathogenesis. Knowledge of the population distribution of MICA alleles and their linkage disequilibrium (LD) with class I human leukocyte antigen (HLA) will enhance our understanding of the potential functional significance of the MICA polymorphism. In the present study, we characterized the MICA and HLA-B polymorphisms in two North American populations: European and African. The individual racial groups showed rather limited variation at the MICA locus, where the same set of three most common alleles, MICA*00201, *004, and *00801, account for 64 and 71% of the allele frequency in European-Americans and African-Americans, respectively. Other common alleles (allele frequency >5% in a population) include MICA*00901 and *010. MICA alleles showed strong linkage disequilibrium with HLA-B. Typically, a common MICA allele has strong LD with several HLA-B alleles, whereas most HLA-B alleles and their related serological groups are associated with a single MICA allele. The lack of evidence for an active diversification of the MICA gene after racial separation indicates an evolutionary history distinct from that of the classical HLA genes.     

Evidence for an inflammatory bowel disease locus on chromosome 3p26: linkage,transmission/disequilibrium and partitioning of linkage

Crohn's disease and ulcerative colitis, the two major forms of idiopathic inflammatory bowel disease (IBD), are heritable, complex traits that appear to share some but not all susceptibility loci. We report that transmission/disequilibrium test analysis of a Crohn's disease genome scan dataset has detected an inflammatory bowel disease locus on chromosome 3p26 (nominal P=0.000052 and genome-wide corrected P=0.039 at D3S1297). An allele sharing method shows significant linkage (multipoint lod=3.69) in a larger, independent sample of inflammatory bowel disease-affected sibling pairs. A survey of 16 chromosome 3p26 short tandem repeat polymorphisms in a combined sample of 234 independent nuclear families with 324 IBD-affected sibling pairs shows significant linkage to chromosome 3p26 (multipoint lod=3.78) and significant transmission/disequilibrium test results at two adjacent markers (nominal P values in two different transmission/disequilibrium analysis methods=0.00011 and 0.0011 for the first marker, and 0.00071 and 0.00013 for the second marker). There is highly significant under-transmission of a common allele and modest over-transmission of other alleles at both markers. Families with no transmission to affected individuals of the under-transmitted alleles show significant linkage (multipoint lod=4.50) that is significantly greater in four simulation studies (P=0.0001, 0.0000625, 0.0000625 and 0.0000625, respectively) than the linkage evidence in families with transmission of the under-transmitted alleles (multipoint lod=0.12). Thus, the existence of an inflammatory bowel disease locus on chromosome 3p26 is supported by significant linkage, transmission/disequilibrium and partitioning of linkage evidence.     

Similarity of the allele frequency and linkage disequilibrium pattern of single nucleotide polymorphisms in drug-related gene loci between Thai and northern East Asian populations: implications for tagging SNP selection in Thais

The similarity of the marker allele frequency and linkage disequilibrium structure between two populations are major factors for the determination of the transferability and efficiency of haplotype tagging SNP derived from one population to use for an indirect association study in another population. To prove the similarity between northern East Asian populations in Hapmap and Thais, 861 SNP in 166 drug-related genes shared between Thais, Han Chinese and Japanese were analyzed for their correlation statistics. Allele frequency, Fst statistics and linkage disequilibrium statistics (r ²) showed a high correlation between these populations. TagSNP sets derived by an aggressive tagging algorithm from these 861 SNP in Japanese and Chinese were used to test the coverage of East Asia-derived tagSNP in Thais. TagSNP derived from Japanese and Chinese are comparable in the percentage of coverage of the alleles captured with tagSNP at r ²≥0.8 (93% vs. 93%) in these drug-related gene loci. Additional tagSNP sets derived from the combination of Japanese- and Chinese-derived tagging SNP sets were used to test the coverage in Thais. The later set improved the percentage of coverage of alleles captured with tagSNP at r ²≥0.8–98% for these sites. High similarity between Thais and northern East Asian allele frequency and linkage disequilibrium statistics supported that tagSNPs derived from the northern East Asian population should be useful for an indirect association study in Thais. The combination of non-overlapping Japanese derived tagSNP and Chinese-derived tagSNP improved the percentage of genomic coverage in Thais, at least in these drug-related gene loci.     

Estimation of haplotype frequency and linkage disequilibrium parameter in the HLA system.

Conventional formulae currently employed were extended to calculate the haplotype frequency and the corresponding variance for an arbitrary number (n) of loci when unrelated individuals were investigated. A formula is presented for the variance for the degree of three locus linkage disequilibrium.     

Evaluating linkage and linkage disequilibrium: use of excess sharing and transmission disequilibrium methods in affected sib pairs

Two popular and robust approaches to analysing affected sib pair (ASP) data for linkage are the traditional excess sharing methods and the transmission/disequilibrium test (TDT). Here we derive an overall test of linkage for multi-allelic ASP marker data which comprises two component tests: one for excess sharing and one for transmission disequilibrium. This method has several advantages. Firstly the overall test of linkage is often more powerful than either of the two component tests. Secondly the method makes it possible to determine the contribution of linkage disequilibrium (LD), in addition to linkage, to an overall positive linkage result. This is useful because the presence of LD in addition to linkage may suggest that the marker locus is in very close proximity to a disease susceptibility gene. Thirdly the method provides estimates of the risk associated with transmission of the different marker alleles.     

X-chromosome as a marker for population history: linkage disequilibrium and haplotype study in Eurasian populations

Linkage disequilibrium (LD) structure is still unpredictable because the interplay of regional recombination rate and demographic history is poorly understood. We have compared the distribution of LD across two genomic regions differing in crossing-over activity -- Xq13 (0.166 cM/Mb) and Xp22 (1.3 cM/Mb) -- in 15 Eurasian populations. Demographic events predicted to increase the LD level -- genetic drift, bottleneck and admixture - had a very strong impact on extent and patterns of regional LD across Xq13 compared to Xp22. The haplotype distribution of the DXS1225-DXS8082 microsatellites from Xq13 exhibiting strong association in all populations was remarkably influenced by population history. European populations shared one common haplotype with a frequency of 25-40%. The Volga-Ural populations studied, living at the geographic borderline of Europe, showed elevated LD as well as harboring a significant fraction of haplotypes originating from East Asia, thus reflecting their past migrations and admixture. In the young Kuusamo isolate from Finland, a bottleneck has led to allelic associations between loci and shifted the haplotype distribution, but has much less affected single microsatellite allele frequencies compared to the main Finnish population. The data show that the footprint of a demographic event is longer preserved in haplotype distribution within a region of low crossing-over rate, than in the information content of a single marker, or between actively recombining markers. As the knowledge of LD patterns is often chosen to assist association mapping of common disease, our conclusions emphasize the importance of understanding the history, structure and variation of a study population.     

High frequency of QPY allele and linkage disequilibrium of granzyme-B in Epstein-Barr-virus-associated hemophagocytic lymphohistiocytosis

Mediation of Epstein-Barr virus (EBV)-specific cytotoxicity in T lymphocyte via the perforin/granzyme pathway has been demonstrated; therefore, a study involving cytolytic molecules was essential for the clarification of hemophagocytic lymphohistiocytosis (HLH) pathogenesis. This investigation, which analysed the frequency of three allelic mutations of granzyme-B (55Q/R, 95P/A and 247Y/H) in patients with EBV-HLH and infectious mononucleosis, identified the high prevalence of the QPY haplotype in EBV-HLH patients in comparison with healthy controls. A > G polymorphism was also detected in intron 5; furthermore, nearly complete linkage disequilibrium was observed among these polymorphisms. The recessive role of the QPY haplotype of granzyme-B might be responsible for the pathogenesis of EBV-HLH. Cytotoxicity and DNA fragmentation of cytotoxic T lymphocytes did not differ among patients characterized by the QPY/QPY, RAH/RAH and QPY/RAH genotypes. This finding suggested that DNA fragmentation in target cells is mediated not only by granzyme-B but also by other molecules, including other granzymes or Fas.     

Evidence of genetic overlap of schizophrenia and bipolar disorder: linkage disequilibrium analysis of chromosome 18 in the Costa Rican population.

The long-standing concept that schizophrenia (SC) and bipolar disorder (BP) represent two distinct illnesses has been recently challenged by findings of overlap of genetic susceptibility loci for these two diseases. We report here the results of a linkage disequilibrium (LD) analysis of chromosome 18 utilizing subjects with SC from the Central Valley of Costa Rica. Evidence of association (P < 0.05) was obtained in three chromosomal regions: 18p11.31 (D18S63), 18q12.3 (D18S474), and 18q22.3-qter (D18S1161, D18S70), all of which overlap or are in close proximity with loci previously shown to be in LD with BP, type I in this population. Since both the SC and bipolar samples contained cases with a history of mania and almost all cases of SC and BP had a history of psychosis, we performed an alternative phenotyping strategy to determine whether presence or absence of mania, in the context of psychosis, would yield distinct linkage patterns along chromosome 18. To address this issue, a cohort of psychotic patients (including a range of DSMIV diagnoses) was divided into two groups based on the presence or absence of mania. Regions that showed association with SC showed segregation of association when the sample was stratified by history of mania. Our results are compared with previous genetic studies of susceptibility to SC or BP, in Costa Rica as well as in other populations. This study illustrates the importance of detailed phenotype analysis in the search for susceptibility genes influencing complex psychiatric disorders in isolated populations and suggests that subdivision of psychoses by presence or absence of past mania syndromes may be useful to define genetic subtypes of chronic psychotic illness.     

Comparative high-resolution analysis of linkage disequilibrium and tag single nucleotide polymorphisms between populations in the vitamin D receptor gene

A genome-wide map of single nucleotide polymorphisms (SNP) and a pattern of linkage disequilibrium (LD) between their alleles are being established in three main ethnic groups. An important question is the applicability of such maps to different populations within a main ethnic group. Therefore, we have developed high-resolution SNP, haplotype and LD maps of vitamin D receptor gene region in large samples from five populations. Comparative analysis reveals that the LD patterns are identical in all four European populations tested with two small regions of 1.3 and 5.7 kb at which LD is disrupted completely resulting in three block-like regions over which there is significant and extensive LD. In an African population the pattern is similar, but two additional LD-breaking spots are also apparent. This LD pattern suggests combined action of recombination hotspots and founder effects, but cannot be explained by random recombination and genetic drift alone. Direct comparison indicates that the tag SNPs selected in one European population effectively predict the non-tag SNPs in the other Europeans, but not in the Gambians, for this region.     

Complete analysis of HLA-DQB1 polymorphism and DR-DQ linkage disequilibrium by oligonucleotide typing

HLA class II polymorphism is functionally important in the control of immune responses, in transplantation immunology, and in the suceptibility to autoimmune diseases. HLA-DQA1 and -DQB1 genes exhibit a larger degree of allelic polymorphism than usually recognized by routine serology. We have therefore performed an extensive analysis of DQB1 polymorphism by oligotyping. A set of 12 oligo probes was hybridized on polymerase chain reaction-amplified DNA, thus allowing the detection of 12 DQB1 alleles, as demonstrated in homozygous as well as in heterozygous individuals. This highly sensitive detection system is particularly relevant within the DQw1 specificity where the 7 allelic sequences can easily be identified. The DQ-DR linkage disequilibrium was analyzed by oligotyping of 80 Caucasoid heterozygous individuals (160 haplotypes), and very tight associations were observed between DRB1 and DQB1 alleles. Five DRB1 alleles, DR-BON, DR4/Dw4 or Dw14, DR7, DRw8.3, and DRw11, however, can be associated with different DQB1 alleles. Moreover the DRB1 and DQB1 oligotyping analysis performed on 20 randomly chosen DRw8 Caucasoid individuals showed a high prevalence of the DRB1*0801-DQB1*0402 haplotype. By combining the analysis of allelic variations at DRB1, DRB3, and DQB1 loci, we can detect 33 different DR-DQ combinations in our panel of Caucasoid individuals. We now apply DQB1 oligotyping on a routine basis for optimal matching of unrelated donors for bone marrow transplantation.     

Inference from the relationships between linkage disequilibrium and allele frequency distributions of 240 candidate SNPs in 109 drug-related genes in four Asian populations

The extensive nucleotide diversity in drug-related genes predisposes individuals to different drug responses and is a major problem in current clinical practice and drug development. Striking allelic frequency differences exist in these genes between populations. In this study, we genotyped 240 sites known to be polymorphic in the Japanese population in each of 270 unrelated healthy individuals comprising 90 each of Malaysian Malays, Indians, and Chinese. These sites are distributed in 109 genes that are drug related, such as genes encoding drug-metabolizing enzymes and drug transporters. Allele frequency and linkage disequilibrium distributions of these sites were determined and compared. They were also compared with similar data of 752 Japanese. Extensive similarities in allele frequency and linkage disequilibrium distributions were observed among Japanese, Malaysian Chinese, and Malays. However, significant differences were observed between Japanese and Malaysian Chinese with Malaysian Indians. These four populations were grouped into two genetic clusters of different ancestries. However, a higher correlation was found between Malaysian Malays and Indians, indicating the existence of extensive admixture between them. The results also imply the possible and rational use of existing single nucleotide polymorphism databases as references to assist future pharmacogenetic studies involving populations of similar ancestry.     

Absence of linkage and linkage disequilibrium to chromosome 15q11-q13 markers in 139 multiplex families with autism

Chromosomal region 15q11-q13 has been implicated to harbor a susceptibility gene or genes underlying autism. Evidence has been derived from the existence of cytogenetic anomalies in this region associated with autism, and the report of linkage in a modest collection of multiplex families. Most recently, linkage disequilibrium with the marker GABRB3-155CA2 in the candidate locus GABRB3, located in this region, has been reported. We searched for linkage using eight microsatellite markers located in this region of chromosome 15 in 147 affected sib-pairs from 139 multiplex autism families. We also tested for linkage disequilibrium in the same set of families with the same markers. We found no evidence for excess allele sharing (linkage) for the markers in this region. Also, we found no evidence of linkage disequilibrium, including for the locus GABRB3-155CA2. Thus, it appears that the role of this region of chromosome 15 is minor, at best, in the majority of individuals with autism. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 88:551–556, 1999. © 1999 Wiley-Liss, Inc.     

The interval of linkage disequilibrium (LD) detected with microsatellite and SNP markers in chromosomes of Finnish populations with different histories

Linkage disequilibrium (LD) has been an efficient tool for fine mapping of monogenic disease genes in population isolates. Its usefulness for identification of predisposing loci for common, polygenic diseases has been challenged on the basis of anticipated allelic and locus heterogeneity. We compared the extent of LD among marker loci in Finnish subpopulations with divergent but well-characterized histories. One study sample represents the early settlement Finnish population, descended from two immigration events 4,000 and 2,000 years ago. The second sample represents the geographically large late settlement region, populated 15 generations ago by several small immigrant groups from the early settlement region. The third is a restricted regional subpopulation in northeastern Finland which was founded 12 generations ago by 39 immigrant families from the late settlement region. We genotyped 243 microsatellite markers and 68 single nucleotide polymorphisms (SNPs) on chromosomes 1q and 5q. The genealogy of the families from the early (n=16) and late settlements (n=54) and the isolated settlement (n=54) was studied in detail back to the 1800s. Microsatellite data revealed greater LD in the young, founder subpopulation than was seen in either of the older populations. Observed linkage disequilibrium correlated not only with physical distance between markers but also with the information content of the markers. Using biallelic SNP markers, significant LD could only be detected up to 0.1 cM. Our results demonstrate the complexity of the concept of 'detectable LD' and emphasize the importance of understanding population history when designing a strategy for disease gene mapping.