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Background Short tandem repeats (STRs) compose approximately 3% of the genome, and mutations at STR loci have been linked to dozens of human diseases including amyotrophic lateral sclerosis, Friedreich ataxia, Huntington disease, and fragile X syndrome. Improving our understanding of these mutations would increase our knowledge of the mutational dynamics of the genome and may uncover additional loci that contribute to disease. To estimate the genome-wide pattern of mutations at STR loci, we analyze blood-derived whole-genome sequencing data for 544 individuals from 29 three-generation CEPH pedigrees. These pedigrees contain both sets of grandparents, the parents, and an average of 9 grandchildren per family. Results We use HipSTR to identify de novo STR mutations in the 2nd generation of these pedigrees and require transmission to the third generation for validation. Analyzing approximately 1.6 million STR loci, we estimate the empirical de novo STR mutation rate to be 5.24 × 10−5 mutations per locus per generation. Perfect repeats mutate about 2 × more often than imperfect repeats. De novo STRs are significantly enriched in Alu elements. Conclusions Approximately 30% of new STR mutations occur within Alu elements, which compose only 11% of the genome, but only 10% are found in LINE-1 insertions, which compose 17% of the genome. Phasing these mutations to the parent of origin shows that parental transmission biases vary among families. We estimate the average number of de novo genome-wide STR mutations per individual to be approximately 85, which is similar to the average number of observed de novo single nucleotide variants.

Germline mutation rates in humans have been estimated for a variety of mutation types, including single-nucleotide and large structural variants. Here, we directly measure the germline retrotransposition rate for the three active retrotransposon elements: L1, Alu, and SVA. We used three tools for calling mobile element insertions (MEIs) (MELT, RUFUS, and TranSurVeyor) on blood-derived whole-genome sequence (WGS) data from 599 CEPH individuals, comprising 33 three-generation pedigrees. We identified 26 de novo MEIs in 437 births. The retrotransposition rate estimates for Alu elements, one in 40 births, is roughly half the rate estimated using phylogenetic analyses, a difference in magnitude similar to that observed for single-nucleotide variants. The L1 retrotransposition rate is one in 63 births and is within range of previous estimates (1:20–1:200 births). The SVA retrotransposition rate, one in 63 births, is much higher than the previous estimate of one in 900 births. Our large, three-generation pedigrees allowed us to assess parent-of-origin effects and the timing of insertion events in either gametogenesis or early embryonic development. We find a statistically significant paternal bias in Alu retrotransposition. Our study represents the first in-depth analysis of the rate and dynamics of human retrotransposition from WGS data in three-generation human pedigrees.

There is strong evidence for a genetic contribution to non-syndromic congenital heart defects (CHDs). However, exome- and genome-wide studies conducted at the variant and gene-level have identified few genome-wide significant CHD-related genes. Gene-set analyses are a useful complement to such studies and candidate gene-set analyses of rare variants have provided insight into the genetics of CHDs. However, similar analyses have not been conducted using data on common genetic variants. Consequently, we conducted common variant analyses of 15 CHD candidate gene-sets, using data from two common types of CHDs: conotruncal heart defects (1431 cases) and left ventricular outflow tract defects (509 cases). After Bonferroni correction for evaluation of multiple gene-sets, the cytoskeletal gene-set was significantly associated with conotruncal heart defects (βS = 0.09, 95% confidence interval (CI) 0.03–0.15). This association was stronger when analyses were restricted to the sub-set of cytoskeletal genes that have been observed to harbor rare damaging genotypes in at least two CHD cases (βS = 0.32, 95% CI 0.08–0.56). These findings add to the evidence linking cytoskeletal genes to CHDs and suggest that, for cytoskeletal genes, common variation may contribute to the risk of CHDs.

Ongoing retrotransposition of Alu, LINE-1, and SINE–VNTR–Alu elements generates diversity and variation among human populations. Previous analyses investigating the population genetics of mobile element insertions (MEIs) have been limited by population ascertainment bias or by relatively small numbers of populations and low sequencing coverage. Here, we use 296 individuals representing 142 global populations from the Simons Genome Diversity Project (SGDP) to discover and characterize MEI diversity from deeply sequenced whole-genome data. We report 5,742 MEIs not originally reported by the 1000 Genomes Project and show that high sampling diversity leads to a 4- to 7-fold increase in MEI discovery rates over the original 1000 Genomes Project data. As a result of negative selection, nonreference polymorphic MEIs are underrepresented within genes, and MEIs within genes are often found in the transcriptional orientation opposite that of the gene. Globally, 80% of Alu subfamilies predate the expansion of modern humans from Africa. Polymorphic MEIs show heterozygosity gradients that decrease from Africa to Eurasia to the Americas, and the number of MEIs found uniquely in a single individual are also distributed in this general pattern. The maximum fraction of MEI diversity partitioned among the seven major SGDP population groups (FST) is 7.4%, similar to, but slightly lower than, previous estimates and likely attributable to the diverse sampling strategy of the SGDP. Finally, we utilize these MEIs to extrapolate the primary Native American shared ancestry component to back to Asia and provide new evidence from genome-wide identical-by-descent genetic markers that add additional support for a southeastern Siberian origin for most Native Americans.

Congenital heart disease (CHD) is the leading cause of mortality from birth defects. Here, exome sequencing of a single cohort of 2,871 CHD probands, including 2,645 parent-offspring trios, implicated rare inherited mutations in 1.8%, including a recessive founder mutation in GDF1 accounting for ∼5% of severe CHD in Ashkenazim, recessive genotypes in MYH6 accounting for ∼11% of Shone complex, and dominant FLT4 mutations accounting for 2.3% of Tetralogy of Fallot. De novo mutations (DNMs) accounted for 8% of cases, including ∼3% of isolated CHD patients and ∼28% with both neurodevelopmental and extra-cardiac congenital anomalies. Seven genes surpassed thresholds for genome-wide significance, and 12 genes not previously implicated in CHD had >70% probability of being disease related. DNMs in ∼440 genes were inferred to contribute to CHD. Striking overlap between genes with damaging DNMs in probands with CHD and autism was also found.

The genetic architecture of sporadic congenital heart disease (CHD) is characterized by enrichment in damaging de novo variants in chromatin-modifying genes. To test the hypothesis that gene pathways contributing to de novo forms of CHD are distinct from those for recessive forms, we analyze 2391 whole-exome trios from the Pediatric Cardiac Genomics Consortium. We deploy a permutation-based gene-burden analysis to identify damaging recessive and compound heterozygous genotypes and disease genes, controlling for confounding effects, such as background mutation rate and ancestry. Cilia-related genes are significantly enriched for damaging rare recessive genotypes, but comparatively depleted for de novo variants. The opposite trend is observed for chromatin-modifying genes. Other cardiac developmental gene classes have less stratification by mode of inheritance than cilia and chromatin-modifying gene classes. Our analyses reveal dominant and recessive CHD are associated with distinct gene functions, with cilia-related genes providing a reservoir of rare segregating variation leading to CHD., Large whole-exome sequencing studies have suggested that the genetic architecture of syndromic congenital heart disease (CHD) is different from sporadic forms. Here, Watkins et al. estimate the relative contribution of damaging recessive and de novo genotypes to CHD in 2391 trios and find them to be associated with different gene functions.

Alu retrotransposons account for more than 10% of the human genome, and insertions of these elements create structural variants segregating in human populations. Such polymorphic Alu are powerful markers to understand population structure, and they represent variants that can greatly impact genome function, including gene expression. Accurate genotyping of Alu and other mobile elements has been challenging. Indeed, we found that Alu genotypes previously called for the 1000 Genomes Project are sometimes erroneous, which poses significant problems for phasing these insertions with other variants that comprise the haplotype. To ameliorate this issue, we introduce a new pipeline -- TypeTE -- which genotypes Alu insertions from whole-genome sequencing data. Starting from a list of polymorphic Alus, TypeTE identifies the hallmarks (poly-A tail and target site duplication) and orientation of Alu insertions using local re-assembly to reconstruct presence and absence alleles. Genotype likelihoods are then computed after re-mapping sequencing reads to the reconstructed alleles. Using a ‘gold standard’ set of PCR-based genotyping of >200 loci, we show that TypeTE improves genotype accuracy from 83% to 92% in the 1000 Genomes dataset. TypeTE can be readily adapted to other retrotransposon families and brings a valuable toolbox addition for population genomics.

Background Polymorphic human Alu elements are excellent tools for assessing population structure, and new retrotransposition events can contribute to disease. Next-generation sequencing has greatly increased the potential to discover Alu elements in human populations, and various sequencing and bioinformatics methods have been designed to tackle the problem of detecting these highly repetitive elements. However, current techniques for Alu discovery may miss rare, polymorphic Alu elements. Combining multiple discovery approaches may provide a better profile of the polymorphic Alu mobilome. AluYb8/9 elements have been a focus of our recent studies as they are young subfamilies (~2.3 million years old) that contribute ~30% of recent polymorphic Alu retrotransposition events. Here, we update our ME-Scan methods for detecting Alu elements and apply these methods to discover new insertions in a large set of individuals with diverse ancestral backgrounds. Results We identified 5,288 putative Alu insertion events, including several hundred novel AluYb8/9 elements from 213 individuals from 18 diverse human populations. Hundreds of these loci were specific to continental populations, and 23 non-reference population-specific loci were validated by PCR. We provide high-quality sequence information for 68 rare AluYb8/9 elements, of which 11 have hallmarks of an active source element. Our subfamily distribution of rare AluYb8/9 elements is consistent with previous datasets, and may be representative of rare loci. We also find that while ME-Scan and low-coverage, whole-genome sequencing (WGS) detect different Alu elements in 41 1000 Genomes individuals, the two methods yield similar population structure results. Conclusion Current in-silico methods for Alu discovery may miss rare, polymorphic Alu elements. Therefore, using multiple techniques can provide a more accurate profile of Alu elements in individuals and populations. We improved our false-negative rate as an indicator of sample quality for future ME-Scan experiments. In conclusion, we demonstrate that ME-Scan is a good supplement for next-generation sequencing methods and is well-suited for population-level analyses.

Here we report the Simons Genome Diversity Project data set: high quality genomes from 300 individuals from 142 diverse populations. These genomes include at least 5.8 million base pairs that are not present in the human reference genome. Our analysis reveals key features of the landscape of human genome variation, including that the rate of accumulation of mutations has accelerated by about 5% in non-Africans compared to Africans since divergence. We show that the ancestors of some pairs of present-day human populations were substantially separated by 100,000 years ago, well before the archaeologically attested onset of behavioural modernity. We also demonstrate that indigenous Australians, New Guineans and Andamanese do not derive substantial ancestry from an early dispersal of modern humans; instead, their modern human ancestry is consistent with coming from the same source as that of other non-Africans., Facultad de Ciencias Naturales y Museo

Inflammatory bowel disease 5 (IBD5) is a 250 kb haplotype on chromosome 5 that is associated with an increased risk of Crohn’s disease in Europeans. The OCTN1 gene is centrally located on IBD5 and encodes a transporter of the antioxidant ergothioneine (ET). The 503F variant of OCTN1 is strongly associated with IBD5 and is a gain-of-function mutation that increases absorption of ET. Although 503F has been implicated as the variant potentially responsible for Crohn’s disease susceptibility at IBD5, there is little evidence beyond statistical association to support its role in disease causation. We hypothesize that 503F is a recent adaptation in Europeans that swept to relatively high frequency and that disease association at IBD5 results not from 503F itself, but from one or more nearby hitchhiking variants, in the genes IRF1 or IL5. To test for evidence of recent positive selection on the 503F allele, we employed the iHS statistic, which was significant in the European CEU HapMap population (P = 0.0007) and European Human Genome Diversity Panel populations (P ≤ 0.01). To evaluate the hypothesis of disease-variant hitchhiking, we performed haplotype association tests on high-density microarray data in a sample of 1,868 Crohn’s disease cases and 5,550 controls. We found that 503F haplotypes with recombination breakpoints between OCTN1 and IRF1 or IL5 were not associated with disease (odds ratio [OR]: 1.05, P = 0.21). In contrast, we observed strong disease association for 503F haplotypes with no recombination between these three genes (OR: 1.24, P = 2.6 × 10−8), as expected if the sweeping haplotype harbored one or more disease-causing mutations in IRF1 or IL5. To further evaluate these disease-gene candidates, we obtained expression data from lower gastrointestinal biopsies of healthy individuals and Crohn’s disease patients. We observed a 72% increase in gene expression of IRF1 among Crohn’s disease patients (P = 0.0006) and no significant difference in expression of OCTN1. Collectively, these data indicate that the 503F variant has increased in frequency due to recent positive selection and that disease-causing variants in linkage disequilibrium with 503F have hitchhiked to relatively high frequency, thus forming the IBD5 risk haplotype. Finally, our association results and expression data support IRF1 as a strong candidate for Crohn’s disease causation.

Much remains unknown about the genetic factors that contribute to essential hypertension. The Utah Genetic Reference Project (UGRP) large pedigree collection provides new opportunities to study quantitative relationships between genetic variation, endophenotypes, and blood pressure.We analyzed the relationship between common single-nucleotide polymorphisms (SNPs) and haplotypes spanning the angiotensinogen (AGT) gene and promoter region, plasma AGT levels, and systolic (SBP) and diastolic blood pressure (DBP) in 424 individuals from 41 two-generation UGRP families.Plasma AGT levels are significantly correlated among UGRP family members. Correlations are higher for males than for females. Parent-offspring correlations for plasma AGT (0.30) are higher than those for SBP (0.26) and DBP (0.17) (all P values0.01). The additive heritability (h(2)) for plasma AGT is high (0.74) and substantially exceeds heritability estimates for SBP (0.26) and DBP (0.16) (all P values0.01). Significant linkage (logarithm of the odds (LOD)3) is found between six AGT SNPs and plasma AGT. A model that utilizes three AGT haplotype groups produces the best LOD score (5.1) that exceeds the best single SNP LOD score (3.8). Plasma AGT and blood pressure were not significantly correlated.Plasma AGT levels demonstrate high heritability in 41 UGRP families. Locus-specific heritability estimates for AGT SNPs and haplotypes approach 67%, indicating that variation at AGT accounts for a large percentage of the heritability of plasma AGT. A three-way haplotype model outperforms single SNPs for quantitative linkage analysis to plasma AGT. In these predominantly normotensive individuals, plasma AGT did not correlate significantly with blood pressure.

We report an analysis of more than 240,000 loci genotyped using the Affymetrix SNP microarray in 554 individuals from 27 worldwide populations in Africa, Asia, and Europe. To provide a more extensive and complete sampling of human genetic variation, we have included caste and tribal samples from two states in South India, Daghestanis from eastern Europe, and the Iban from Malaysia. Consistent with observations made by Charles Darwin, our results highlight shared variation among human populations and demonstrate that much genetic variation is geographically continuous. At the same time, principal components analyses reveal discernible genetic differentiation among almost all identified populations in our sample, and in most cases, individuals can be clearly assigned to defined populations on the basis of SNP genotypes. All individuals are accurately classified into continental groups using a model-based clustering algorithm, but between closely related populations, genetic and self-classifications conflict for some individuals. The 250K data permitted high-level resolution of genetic variation among Indian caste and tribal populations and between highland and lowland Daghestani populations. In particular, upper-caste individuals from Tamil Nadu and Andhra Pradesh form one defined group, lower-caste individuals from these two states form another, and the tribal Irula samples form a third. Our results emphasize the correlation of genetic and geographic distances and highlight other elements, including social factors that have contributed to population structure.

The study of ethnically homogeneous populations may help to identify schizophrenia risk loci. The authors conducted a genomewide linkage scan for schizophrenia in an Indian population.Participants were 441 individuals (262 affected probands and siblings) who were recruited primarily from one ethnically homogeneous group, the Tamil Brahmin caste, although individuals from other geographically proximal castes also participated. Genotyping of 124 affected sibling pair pedigrees was performed with 402 short tandem repeat polymorphisms. Linkage analyses were conducted using nonparametric exponential LOD (logarithm of the odds ratio for linkage) scores and parametric heterogeneity LOD scores. Parametric heterogeneity scores were calculated using simple dominant and recessive models, correcting for multiple statistics. The data were examined for evidence of consanguinity. Genomewide significance levels were determined using 10,000 gene dropping simulations.These findings revealed genomewide significant linkage to chromosome 1p31.1, through the use of both exponential and heterogeneity LOD scores, incorporating correction for multiple statistics and mild consanguinity. The estimated sibling recurrence risk associated with this putative locus was 1.95. Analysis for heterogeneity LOD scores also detected suggestive linkage to chromosomes 13q22.1 and 16q12.2. Using 117 tag single nucleotide polymorphisms (SNPs), family-based association analyses of phosphodiesterase 4B (PDE4B), the closest schizophrenia candidate gene, detected no convincing evidence of association, suggesting that the chromosome 1 peak represents a novel risk locus.This is the first study-to the authors' knowledge-to report significant linkage of schizophrenia to chromosome 1p31.1. Further investigation of this chromosome region in diverse populations is warranted to identify underlying sequence variants.

Current microarray technology allows researchers to genotype a large number of SNPs with relatively small amounts of DNA. Nevertheless, researchers and clinicians still frequently face the problem of acquiring enough high-quality DNA for analysis. Whole-genome amplification (WGA) methods offer a solution for this problem, and earlier studies have shown that WGA samples perform reasonably well in small-scale genetic analyses (e.g. Affymetrix 10K array). To determine the performance of WGA products on a large-scale genotyping array, we compared the Affymetrix 250K array genotyping results of genomic DNA and their WGA products from four individuals. Our results indicate that WGA product performs well on the 250K array compared to genomic DNA, especially when using the BRLMM calling algorithm. WGA samples have high call rates (97.5% on average, compared to 99.4% for genomic DNA) and excellent concordance rates with their corresponding genomic DNA samples (98.7% on average). In addition, no apparent systematic genomic amplification bias can be detected. This study demonstrates that, although there is a slight decrease in the total call rates, WGA methods provide a reliable approach for increasing the amount of DNA samples for use with a common SNP genotyping array.

The ETS gene family is frequently involved in chromosome translocations that cause human cancer, including prostate cancer, leukemia, and sarcoma. However, the mechanisms by which oncogenic ETS proteins, which are DNA-binding transcription factors, target genes necessary for tumorigenesis is not well understood. Ewing's sarcoma serves as a paradigm for the entire class of ETS-associated tumors because nearly all cases harbor recurrent chromosomal translocations involving ETS genes. The most common translocation in Ewing's sarcoma encodes the EWS/FLI oncogenic transcription factor. We used whole genome localization (ChIP-chip) to identify target genes that are directly bound by EWS/FLI. Analysis of the promoters of these genes demonstrated a significant over-representation of highly repetitive GGAA-containing elements (microsatellites). In a parallel approach, we found that EWS/FLI uses GGAA microsatellites to regulate the expression of some of its target genes including NR0B1 , a gene required for Ewing's sarcoma oncogenesis. The microsatellite in the NR0B1 promoter bound EWS/FLI in vitro and in vivo and was both necessary and sufficient to confer EWS/FLI regulation to a reporter gene. Genome wide computational studies demonstrated that GGAA microsatellites were enriched close to EWS/FLI-up-regulated genes but not down-regulated genes. Mechanistic studies demonstrated that the ability of EWS/FLI to bind DNA and modulate gene expression through these repetitive elements depended on the number of consecutive GGAA motifs. These findings illustrate an unprecedented route to specificity for ETS proteins and use of microsatellites in tumorigenesis.

Sudmant, Peter H. et al., In order to explore the diversity and selective signatures of duplication and deletion human copy-number variants (CNVs), we sequenced 236 individuals from 125 distinct human populations. We observed that duplications exhibit fundamentally different population genetic and selective signatures than deletions and are more likely to be stratified between human populations. Through reconstruction of the ancestral human genome, we identify megabases of DNA lost in different human lineages and pinpoint large duplications that introgressed from the extinct Denisova lineage now found at high frequency exclusively in Oceanic populations. We find that the proportion of CNV base pairs to single-nucleotide–variant base pairs is greater among non-Africans than it is among African populations, but we conclude that this difference is likely due to unique aspects of non-African population history as opposed to differences in CNV load., This project has been funded in part with federal funds from the National Cancer Institute, NIH, under contract HHSN26120080001E. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does the mention of trade names, commercial products, or organizations imply endorsement by the U.S. government. This research was supported in part by the Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research. This work was also partly supported by NIH grant 2R01HG002385 and a grant (11631) from the Paul G. Allen Family Foundation to E.E.E. The sequencing for this study was supported by a grant from the Simons Foundation to D.R. (SFARI 280376) and by a HOMINID grant from the NSF to D.R. (BCS-1032255). T.K. is supported by a European Research Council Starting Investigator grant (FP7 - 26213). R.S. and S.D. received support from the Ministry of Education and Science, Russian Federation (14.Z50.31.0010). H.S., E.M., R.V., and M.M. are supported by Institutional Research Funding from the Estonian Research Council IUT24-1 and by the European Regional Development Fund (European Union) through the Centre of Excellence in Genomics to Estonian Biocentre and University of Tartu. S.A.T. is supported by NIH grants 5DP1ES022577 05, 1R01DK104339-01, and 1R01GM113657-01. C.T.-S. is supported by Wellcome Trust grant 098051. C.M.B. is supported by the NSF (award numbers 0924726 and 1153911). E.E.E. and D.R. are investigators of the Howard Hughes Medical Institute. Data are deposited into ENA (PRJEB9586 or ERP010710), and variant calls are deposited in dbVar (PRJNA285786). E.E.E. is on the scientific advisory board of DNAnexus, Incorporated, and is a consultant for Kunming University of Science and Technology (KUST) as part of the 1000 China Talent Program.

Understanding the distribution of human genetic variation is an important foundation for research into the genetics of common diseases. Some of the alleles that modify common disease risk are themselves likely to be common and, thus, amenable to identification using gene-association methods. A problem with this approach is that the large sample sizes required for sufficient statistical power to detect alleles with moderate effect make gene-association studies susceptible to false-positive findings as the result of population stratification 12. Such type I errors can be eliminated by using either family-based association tests or methods that sufficiently adjust for population stratification 345. These methods require the availability of genetic markers that can detect and, thus, control for sources of genetic stratification among populations. In an effort to investigate population stratification and identify appropriate marker panels, we have analysed 11,555 single nucleotide polymorphisms in 203 individuals from 12 diverse human populations. Individuals in each population cluster to the exclusion of individuals from other populations using two clustering methods. Higher-order branching and clustering of the populations are consistent with the geographic origins of populations and with previously published genetic analyses. These data provide a valuable resource for the definition of marker panels to detect and control for population stratification in population-based gene identification studies. Using three US resident populations (European-American, African-American and Puerto Rican), we demonstrate how such studies can proceed, quantifying proportional ancestry levels and detecting significant admixture structure in each of these populations.

The Alu Yb-lineage is a 'young' primarily human-specific group of short interspersed element (SINE) subfamilies that have integrated throughout the human genome. In this study, we have computationally screened the draft sequence of the human genome for Alu Yb-lineage subfamily members present on autosomal chromosomes. A total of 1,733 Yb Alu subfamily members have integrated into human autosomes. The average ages of Yb-lineage subfamilies, Yb7, Yb8 and Yb9, are estimated as 4.81, 2.39 and 2.32 million years, respectively. In order to determine the contribution of the Alu Yb-lineage to human genomic diversity, 1,202 loci were analysed using polymerase chain reaction (PCR)-based assays, which amplify the genomic regions containing individual Yb-lineage subfamily members. Approximately 20 per cent of the Yb-lineage Alu elements are polymorphic for insertion presence/absence in the human genome. Fewer than 0.5 per cent of the Yb loci also demonstrate insertions at orthologous positions in non-human primate genomes. Genomic sequencing of these unusual loci demonstrates that each of the orthologous loci from non-human primate genomes contains older Y, Sg and Sx Alu family members that have been altered, through various mechanisms, into Yb8 sequences. These data suggest that Alu Yb-lineage subfamily members are largely restricted to the human genome. The high copy number, level of insertion polymorphism and estimated age indicate that members of the Alu Yb elements will be useful in a wide range of genetic analyses.

The L1 Ta subfamily of long interspersed elements (LINEs) consists exclusively of human-specific L1 elements. Polymerase chain reaction-based screening in nonhuman primate genomes of the orthologous sites for 249 human L1 Ta elements resulted in the recovery of various types of sequence variants for approximately 12% of these loci. Sequence analysis was employed to capture the nature of the observed variation and to determine the levels of gene conversion and insertion site homoplasy associated with LINE elements. Half of the orthologous loci differed from the predicted sizes due to localized sequence variants that occurred as a result of common mutational processes in ancestral sequences, often including regions containing simple sequence repeats. Additional sequence variation included genomic deletions that occurred upon L1 insertion, as well as successive mobile element insertions that accumulated within a single locus over evolutionary time. Parallel independent mobile element insertions at orthologous loci in distinct species may introduce homoplasy into retroelement-based phylogenetic and population genetic data. We estimate the overall frequency of parallel independent insertion events at L1 insertion sites in seven different primate species to be very low (0.52%). In addition, no cases of insertion site homoplasy involved the integration of a second L1 element at any of the loci, but rather largely involved secondary insertions of Alu elements. No independent mobile element insertion events were found at orthologous loci in the human and chimpanzee genomes. Therefore, L1 insertion polymorphisms appear to be essentially homoplasy free characters well suited for the study of population genetics and phylogenetic relationships within closely related species.

The preTa subfamily of long interspersed elements (LINEs) is characterized by a three base-pair “ACG” sequence in the 3′ untranslated region, contains approximately 400 members in the human genome, and has low level of nucleotide divergence with an estimated average age of 2.34 million years old suggesting that expansion of the L1 preTa subfamily occurred just after the divergence of humans and African apes. We have identified 362 preTa L1 elements from the draft human genomic sequence, investigated the genomic characteristics of preTa L1 insertions, and screened individual elements across diverse human populations and various non-human primate species using polymerase chain reaction (PCR) assays to determine the phylogenetic origin and levels of human genomic diversity associated with the L1 elements. All of the preTa L1 elements analyzed by PCR were absent from the orthologous positions in non-human primate genomes with 33 (14%) of the L1 elements being polymorphic with respect to insertion presence or absence in the human genome. The newly identified L1 insertion polymorphisms will prove useful as identical by descent genetic markers for the study of human population genetics. We provide evidence that preTa L1 elements show an integration site preference for genomic regions with low GC content. Computational analysis of the preTa L1 elements revealed that 29% of the elements amenable to complete sequence analysis have apparently escaped 5′ truncation and are essentially full-length (approximately 6 kb). In all, 29 have two intact open reading frames and may be capable of retrotransposition.

Author(s): Monument, Michael J; Johnson, Kirsten M; McIlvaine, Elizabeth; Abegglen, Lisa; Watkins, W Scott; Jorde, Lynn B; Womer, Richard B; Beeler, Natalie; Monovich, Laura; Lawlor, Elizabeth R; Bridge, Julia A; Schiffman, Joshua D; Krailo, Mark D; Randall, R Lor; Lessnick, Stephen L | Abstract: BackgroundThe genetics involved in Ewing sarcoma susceptibility and prognosis are poorly understood. EWS/FLI and related EWS/ETS chimeras upregulate numerous gene targets via promoter-based GGAA-microsatellite response elements. These microsatellites are highly polymorphic in humans, and preliminary evidence suggests EWS/FLI-mediated gene expression is highly dependent on the number of GGAA motifs within the microsatellite.ObjectivesHere we sought to examine the polymorphic spectrum of a GGAA-microsatellite within the NR0B1 promoter (a critical EWS/FLI target) in primary Ewing sarcoma tumors, and characterize how this polymorphism influences gene expression and clinical outcomes.ResultsA complex, bimodal pattern of EWS/FLI-mediated gene expression was observed across a wide range of GGAA motifs, with maximal expression observed in constructs containing 20-26 GGAA motifs. Relative to white European and African controls, the NR0B1 GGAA-microsatellite in tumor cells demonstrated a strong bias for haplotypes containing 21-25 GGAA motifs suggesting a relationship between microsatellite function and disease susceptibility. This selection bias was not a product of microsatellite instability in tumor samples, nor was there a correlation between NR0B1 GGAA-microsatellite polymorphisms and survival outcomes.ConclusionsThese data suggest that GGAA-microsatellite polymorphisms observed in human populations modulate EWS/FLI-mediated gene expression and may influence disease susceptibility in Ewing sarcoma.

We have utilized computational biology to screen GenBank for the presence of recently integrated Ya5 and Yb8 Alu family members. Our analysis identified 2640 Ya5 Alu family members and 1852 Yb8 Alu family members from the draft sequence of the human genome. We selected a set of 475 of these elements for detailed analyses. Analysis of the DNA sequences from the individual Alu elements revealed a low level of random mutations within both subfamilies consistent with the recent origin of these elements within the human genome. Polymerase chain reaction assays were used to determine the phylogenetic distribution and human genomic variation associated with each Alu repeat. Over 99 % of the Ya5 and Yb8 Alu family members were restricted to the human genome and absent from orthologous positions within the genomes of several non-human primates, confirming the recent origin of these Alu subfamilies in the human genome. Approximately 1 % of the analyzed Ya5 and Yb8 Alu family members had integrated into previously undefined repeated regions of the human genome. Analysis of mosaic Yb8 elements suggests gene conversion played an important role in generating sequence diversity among these elements. Of the 475 evaluated elements, a total of 106 of the Ya5 and Yb8 Alu family members were polymorphic for insertion presence/absence within the genomes of a diverse array of human populations. The newly identified Alu insertion polymorphisms will be useful tools for the study of human genomic diversity.

The origins and affinities of the approximately 1 billion people living on the subcontinent of India have long been contested. This is owing, in part, to the many different waves of immigrants that have influenced the genetic structure of India. In the most recent of these waves, Indo-European-speaking people from West Eurasia entered India from the Northwest and diffused throughout the subcontinent. They purportedly admixed with or displaced indigenous Dravidic-speaking populations. Subsequently they may have established the Hindu caste system and placed themselves primarily in castes of higher rank. To explore the impact of West Eurasians on contemporary Indian caste populations, we compared mtDNA (400 bp of hypervariable region 1 and 14 restriction site polymorphisms) and Y-chromosome (20 biallelic polymorphisms and 5 short tandem repeats) variation in approximately 265 males from eight castes of different rank to approximately 750 Africans, Asians, Europeans, and other Indians. For maternally inherited mtDNA, each caste is most similar to Asians. However, 20%-30% of Indian mtDNA haplotypes belong to West Eurasian haplogroups, and the frequency of these haplotypes is proportional to caste rank, the highest frequency of West Eurasian haplotypes being found in the upper castes. In contrast, for paternally inherited Y-chromosome variation each caste is more similar to Europeans than to Asians. Moreover, the affinity to Europeans is proportionate to caste rank, the upper castes being most similar to Europeans, particularly East Europeans. These findings are consistent with greater West Eurasian male admixture with castes of higher rank. Nevertheless, the mitochondrial genome and the Y chromosome each represents only a single haploid locus and is more susceptible to large stochastic variation, bottlenecks, and selective sweeps. Thus, to increase the power of our analysis, we assayed 40 independent, biparentally inherited autosomal loci (1 LINE-1 and 39 Alu elements) in all of the caste and continental populations (approximately 600 individuals). Analysis of these data demonstrated that the upper castes have a higher affinity to Europeans than to Asians, and the upper castes are significantly more similar to Europeans than are the lower castes. Collectively, all five datasets show a trend toward upper castes being more similar to Europeans, whereas lower castes are more similar to Asians. We conclude that Indian castes are most likely to be of proto-Asian origin with West Eurasian admixture resulting in rank-related and sex-specific differences in the genetic affinities of castes to Asians and Europeans.

Alu retrotransposons are the most numerous and active mobile elements in humans, causing genetic disease and creating genomic diversity. Mobile element scanning (ME-Scan) enables comprehensive and affordable identification of mobile element insertions (MEI) using targeted high-throughput sequencing of multiplexed MEI junction libraries. In a single experiment, ME-Scan identifies nearly all AluYb8 and AluYb9 elements, with high sensitivity for both rare and common insertions, in 169 individuals of diverse ancestry. ME-Scan detects heterozygous insertions in single individuals with 91% sensitivity. Insertion presence or absence states determined by ME-Scan are 95% concordant with those determined by locus-specific PCR assays. By sampling diverse populations from Africa, South Asia, and Europe, we are able to identify 5799 Alu insertions, including 2524 novel ones, some of which occur in exons. Sub-Saharan populations and a Pygmy group in particular carry numerous intermediate-frequency Alu insertions that are absent in non-African groups. There is a significant dearth of exon-interrupting insertions among common Alu polymorphisms, but the density of singleton Alu insertions is constant across exonic and nonexonic regions. In one case, a validated novel singleton Alu interrupts a protein-coding exon of FAM187B. This implies that exonic Alu insertions are generally deleterious and thus eliminated by natural selection, but not so quickly that they cannot be observed as extremely rare variants.

Deedu (DU) Mongolians, who migrated from the Mongolian steppes to the Qinghai-Tibetan Plateau approximately 500 years ago, are challenged by environmental conditions similar to native Tibetan highlanders. Identification of adaptive genetic factors in this population could provide insight into coordinated physiological responses to this environment. Here we examine genomic and phenotypic variation in this unique population and present the first complete analysis of a Mongolian whole-genome sequence. High-density SNP array data demonstrate that DU Mongolians share genetic ancestry with other Mongolian as well as Tibetan populations, specifically in genomic regions related with adaptation to high altitude. Several selection candidate genes identified in DU Mongolians are shared with other Asian groups (e.g., EDAR), neighboring Tibetan populations (including high-altitude candidates EPAS1, PKLR, and CYP2E1), as well as genes previously hypothesized to be associated with metabolic adaptation (e.g., PPARG). Hemoglobin concentration, a trait associated with high-altitude adaptation in Tibetans, is at an intermediate level in DU Mongolians compared to Tibetans and Han Chinese at comparable altitude. Whole-genome sequence from a DU Mongolian (Tianjiao1) shows that about 2% of the genomic variants, including more than 300 protein-coding changes, are specific to this individual. Our analyses of DU Mongolians and the first Mongolian genome provide valuable insight into genetic adaptation to extreme environments., Author Summary Throughout history, Mongolians have survived the harsh conditions of northern latitudes, including seasonal cold, drought, and a restricted diet. Approximately 500 years ago, nomadic Deedu (DU; “at high altitude”) Mongolians migrated from the Mongolian steppes to the northeastern highlands of the Qinghai-Tibetan Plateau. Using high-density SNP data, we demonstrate that present-day DU Mongolians share genetic ancestry with other Mongolians and with Tibetans. High-altitude selection candidate genes previously identified in the latter population (EPAS1, PKLR, CYP2E1), and PPARG, a gene long hypothesized to play a role in metabolic adaptation, are among the strongest adaptive signals in DU Mongolians. Furthermore, we show that hemoglobin concentration, associated with high-altitude adaptation in Tibetans, is intermediate in DU Mongolians compared to Tibetans and Han Chinese at comparable altitudes. Whole-genome sequence from a DU Mongolian shows that approximately 300 protein-coding changes are specific to this individual. Our analyses provide new perspectives on genetic variation and adaptation to extreme environments.

Background Because of the role of inflammation in preterm birth (PTB), polymorphisms in and near the interleukin-6 gene (IL6) have been association study targets. Several previous studies have assessed the association between PTB and a single nucleotide polymorphism (SNP), rs1800795, located in the IL6 gene promoter region. Their results have been inconsistent and SNP frequencies have varied strikingly among different populations. We therefore conducted a meta-analysis with subgroup analysis by population strata to: (1) reduce the confounding effect of population structure, (2) increase sample size and statistical power, and (3) elucidate the association between rs1800975 and PTB. Results We reviewed all published papers for PTB phenotype and SNP rs1800795 genotype. Maternal genotype and fetal genotype were analyzed separately and the analyses were stratified by population. The PTB phenotype was defined as gestational age (GA) < 37 weeks, but results from earlier GA were selected when available. All studies were compared by genotype (CC versus CG+GG), based on functional studies. For the maternal genotype analysis, 1,165 PTBs and 3,830 term controls were evaluated. Populations were stratified into women of European descent (for whom the most data were available) and women of heterogeneous origin or admixed populations. All ancestry was self-reported. Women of European descent had a summary odds ratio (OR) of 0.68, (95% confidence interval (CI) 0.51 – 0.91), indicating that the CC genotype is protective against PTB. The result for non-European women was not statistically significant (OR 1.01, 95% CI 0.59 - 1.75). For the fetal genotype analysis, four studies were included; there was no significant association with PTB (OR 0.98, 95% CI 0.72 - 1.33). Sensitivity analysis showed that preterm premature rupture of membrane (PPROM) may be a confounding factor contributing to phenotype heterogeneity. Conclusions IL6 SNP rs1800795 genotype CC is protective against PTB in women of European descent. It is not significant in other heterogeneous or admixed populations, or in fetal genotype analysis. Population structure is an important confounding factor that should be controlled for in studies of PTB.

Background Populations of the Americas were founded by early migrants from Asia, and some have experienced recent genetic admixture. To better characterize the native and non-native ancestry components in populations from the Americas, we analyzed 815,377 autosomal SNPs, mitochondrial hypervariable segments I and II, and 36 Y-chromosome STRs from 24 Mesoamerican Totonacs and 23 South American Bolivians. Results and Conclusions We analyzed common genomic regions from native Bolivian and Totonac populations to identify 324 highly predictive Native American ancestry informative markers (AIMs). As few as 40–50 of these AIMs perform nearly as well as large panels of random genome-wide SNPs for predicting and estimating Native American ancestry and admixture levels. These AIMs have greater New World vs. Old World specificity than previous AIMs sets. We identify highly-divergent New World SNPs that coincide with high-frequency haplotypes found at similar frequencies in all populations examined, including the HGDP Pima, Maya, Colombian, Karitiana, and Surui American populations. Some of these regions are potential candidates for positive selection. European admixture in the Bolivian sample is approximately 12%, though individual estimates range from 0–48%. We estimate that the admixture occurred ~360–384 years ago. Little evidence of European or African admixture was found in Totonac individuals. Bolivians with pre-Columbian mtDNA and Y-chromosome haplogroups had 5–30% autosomal European ancestry, demonstrating the limitations of Y-chromosome and mtDNA haplogroups and the need for autosomal ancestry informative markers for assessing ancestry in admixed populations.

The genetics of Ewing sarcoma development remain obscure. The incidence of Ewing sarcoma is ten-fold less in Africans as compared to Europeans, irrespective of geographic location, suggesting population-specific genetic influences. Since GGAA-containing microsatellites within key target genes are necessary for Ewing sarcoma–specific EWS/FLI DNA binding and gene activation, and gene expression is positively correlated with the number of repeat motifs in the promoter/enhancer region, we sought to determine if significant polymorphisms exist between African and European populations which might contribute to observed differences in Ewing sarcoma incidence and outcomes. GGAA microsatellites upstream of two critical EWS/FLI target genes, NR0B1 and CAV1, were sequenced from subjects of European and African descent. While the characteristics of the CAV1 promoter microsatellites were similar across both populations, the NR0B1 microsatellite in African subjects was significantly larger, harboring more repeat motifs, a greater number of repeat segments, and longer consecutive repeats, than in European subjects. These results are biologically intriguing as NR0B1 was the most highly enriched EWS/FLI bound gene in prior studies, and is absolutely necessary for oncogenic transformation in Ewing sarcoma. These data suggest that GGAA microsatellite polymorphisms in the NR0B1 gene might influence disease susceptibility and prognosis in Ewing sarcoma in unanticipated ways.

Of the > 50 different genetic variants of human serum albumin (alloalbumins) that have been characterized by amino acid or DNA sequence analysis, almost half have been identified in Italy through a long-term electrophoretic survey of serum. Previously we have reported structural studies of 11 Italian alloalbumins with point mutations, 2 different carboxyl-terminal variants, and 1 case of analbuminemia in an Italian family. This article describes confirmation by DNA sequencing of mutations previously inferred from protein sequencing of 4 of the above alloalbumins; it also reports the mutations identified by protein and DNA sequence analysis of 4 other Italian alloalbumins not previously recorded: albumin Larino, His3-->Tyr; Tradate-2 (protein sequencing only), Lys225-->Gln; Caserta, Lys276-->Asn; and Bazzano, a carboxyl-terminal variant. The first 3 have point mutations that produce a single amino acid substitution, but a nucleotide deletion causes a frameshift and an altered and truncated carboxyl-terminal sequence in albumin Bazzano. In these 4 instances the expression of the alloalbumin is variable, ranging from 10% to 70% of the total albumiN, in contrast to the usual 50% each for the normal and mutant albumin. The distribution of point mutations in the albumin gene is nonrandom; most of the 47 reported point substitutions involve charged amino acid residues on the surface of the molecule that are not concerned with ligand-binding sites.

Accurate estimation of recent shared ancestry is important for genetics, evolution, medicine, conservation biology, and forensics. Established methods estimate kinship accurately for first-degree through third-degree relatives. We demonstrate that chromosomal segments shared by two individuals due to identity by descent (IBD) provide much additional information about shared ancestry. We developed a maximum-likelihood method for the estimation of recent shared ancestry (ERSA) from the number and lengths of IBD segments derived from high-density SNP or whole-genome sequence data. We used ERSA to estimate relationships from SNP genotypes in 169 individuals from three large, well-defined human pedigrees. ERSA is accurate to within one degree of relationship for 97% of first-degree through fifth-degree relatives and 80% of sixth-degree and seventh-degree relatives. We demonstrate that ERSA's statistical power approaches the maximum theoretical limit imposed by the fact that distant relatives frequently share no DNA through a common ancestor. ERSA greatly expands the range of relationships that can be estimated from genetic data and is implemented in a freely available software package.

Heart failure is a leading cause of death of people in South Asia, and cardiomyopathy is a major cause of heart failure. Myosin binding protein C (MYBPC3) is expressed in the heart muscle, where it regulates the cardiac response to adrenergic stimulation and is important for the structural integrity of the sarcomere. Mutations in the MYBPC3 gene are associated with hypertrophic or dilated cardiomyopathies. A 25-base-pair deletion in intron 32 causes skipping of the downstream exon and is associated with familial cardiomyopathy. To date, this deletion is found primarily in India and South Asia, although it is also found at low frequency in Southeast Asia. In order to better characterise the distribution of this variant, we determined its frequency in 447 individuals from 19 populations, including 10 populations from India and neighbouring populations from Pakistan and Nepal. The deletion frequency is over 8% in some of our Indian samples, and it is not present in any of the populations we sampled outside of India. The differences in the deletion frequencies among populations in India are consistent with patterns of variation previously reported and with patterns we observed among Indian populations based on high-density SNP chip data. Our results indicate that the MYBPC3 deletion is primarily found among Indian populations and that its distribution is consistent with genome-wide patterns of variation in India.

Background Recombination rates vary widely across the human genome, but little of that variation is correlated with known DNA sequence features. The genome contains more than one million Alu mobile element insertions, and these insertions have been implicated in non-homologous recombination, modulation of DNA methylation, and transcriptional regulation. If individual Alu insertions have even modest effects on local recombination rates, they could collectively have a significant impact on the pattern of linkage disequilibrium in the human genome and on the evolution of the Alu family itself. Results We carried out sequencing, SNP identification, and SNP genotyping around 19 AluY insertion loci in 347 individuals sampled from diverse populations, then used the SNP genotypes to estimate local recombination rates around the AluY loci. The loci and SNPs were chosen so as to minimize other factors (such as SNP ascertainment bias and SNP density) that could influence recombination rate estimates. We detected a significant increase in recombination rate within ~2 kb of the AluY insertions in our African population sample. To test this observation against a larger set of AluY insertions, we applied our locus- and SNP-selection design and analyses to the HapMap Phase II data. In that data set, we observed a significantly increased recombination rate near AluY insertions in both the CEU and YRI populations. Conclusion We show that the presence of a fixed AluY insertion is significantly predictive of an elevated local recombination rate within 2 kb of the insertion, independent of other known predictors. The magnitude of this effect, approximately a 6% increase, is comparable to the effects of some recombinogenic DNA sequence motifs identified via their association with recombination hot spots.

Linkage disequilibrium (LD) has received much attention recently because of its value in localizing disease-causing genes. Due to the extensive LD between neighboring loci in the human genome, it is believed that a subset of the single nucleotide polymorphisms in a region (tagSNPs) can be selected to capture most of the remaining SNP variants. In this study, we examined LD patterns and HapMap tagSNP transferability in more than 300 individuals. A South Indian sample and an African Mbuti Pygmy population sample were included to evaluate the performance of HapMap tagSNPs in geographically distinct and genetically isolated populations. Our results show that HapMap tagSNPs selected with r2 >= 0.8 can capture more than 85% of the SNPs in populations that are from the same continental group. Combined tagSNPs from HapMap CEU and CHB+JPT serve as the best reference for the Indian sample. The HapMap YRI are a sufficient reference for tagSNP selection in the Pygmy sample. In addition to our findings, we reviewed over 25 recent studies of tagSNP transferability and propose a general guideline for selecting tagSNPs from HapMap populations.

Autosomal short-tandem repeats (STRs) were typed in ethnic populations of Kubachians, Dargins, Avars, Lezgins, Kumiks, and Nogais of the Caucasus (Daghestan, Russia) at the University of Utah. Daghestan ethnic populations demonstrated differences in STR allele frequency distributions, but these differences were much lower among these ethnic groups compared to worldwide ethnic groups. The observed genetic diversity was low while F(ST) values were high, both of which provided supporting evidence for small population sizes and high levels of isolation among the ethnic groups. An analysis of genetic distance from the three major continents, encompassing Daghestan populations and groups, reveals three distinct clusters: all populations of African affiliation, European and Daghestan populations except the Nogais, and Asian populations with ethnic Nogais.

Genetic, ethnographic, and historical evidence suggests that the Hindu castes have been highly endogamous for several thousand years and that, when movement between castes does occur, it typically consists of females joining castes of higher social status. However, little is known about migration rates in these populations or the extent to which migration occurs between caste groups of low, middle, and high social status. To investigate these aspects of migration, we analyzed the largest collection of genetic markers collected to date in Hindu caste populations. These data included 45 newly typed autosomal short tandem repeat polymorphisms (STRPs), 411 bp of mitochondrial DNA sequence, and 43 Y-chromosomal single-nucleotide polymorphisms that were assayed in more than 200 individuals of known caste status sampled in Andrah Pradesh, in South India. Application of recently developed likelihood-based analyses to this dataset enabled us to obtain genetically derived estimates of intercaste migration rates. STRPs indicated migration rates of 1-2% per generation between high-, middle-, and low-status caste groups. We also found support for the hypothesis that rates of gene flow differ between maternally and paternally inherited genes. Migration rates were substantially higher in maternally than in paternally inherited markers. In addition, while prevailing patterns of migration involved movement between castes of similar rank, paternally inherited markers in the low-status castes were most likely to move into high-status castes. Our findings support earlier evidence that the caste system has been a significant, long-term source of population structuring in South Indian Hindu populations, and that patterns of migration differ between males and females.

Alu elements have inserted in the human genome throughout primate evolution. A small number of Alu insertions have occurred after the divergence of humans from nonhuman primates and therefore should not be present in nonhuman primate genomes. Most of these recently integrated Alu elements are contained with a series of discrete Alu subfamilies that are related to each other based upon diagnostic nucleotide substitutions. We have extracted members of the Alu Yd subfamily that are derivatives of the Alu Y subfamily that share a common 12-bp deletion that defines the Yd lineage from the draft sequence of the human genome. Analysis of the Yd Alu elements resulted in the recovery of two new Alu subfamilies, Yd3 and Yd6, which contain a total of 295 members (198 Yd3 and 97 Yd6). DNA sequence analysis of each of the Alu Yd subfamilies yielded age estimates of 8.02 and 1.20 million years old for the Alu Yd3 and Yd6 subfamilies, respectively. Two hundred Alu Yd3 and Yd6 loci were screened using polymerase chain reaction (PCR) assays to determine their phylogenetic origin and associated levels of human genomic diversity. The Alu Yd3 subfamily appears to have started amplifying relatively early in primate evolution and continued propagating albeit at a low level as many of its members are found in a variety of hominoid (humans, greater and lesser ape) genomes. Only two of the elements are polymorphic in the human genome and absent from the genomes of nonhuman primates. By contrast all of the members of the Alu Yd6 subfamily are restricted to the human genome, with 12% of the elements representing insertion polymorphisms in human populations. A single Alu Yd6 locus contained an independent parallel forward insertion of a paralogous Alu Sq sequence in the owl monkey. These Alu subfamilies are a source of genomic fossil relics for the study of primate phylogenetics and human population genetics.

A comprehensive analysis of the human sex chromosomes was undertaken to assess Alu-associated human genomic diversity and to identify novel Alu insertion polymorphisms for the study of human evolution. Three hundred forty-five recently integrated Alu elements from eight different Alu subfamilies were identified on the X and Y chromosomes, 225 of which were selected and analyzed by polymerase chain reaction (PCR). From a total of 225 elements analyzed, 16 were found to be polymorphic on the X chromosome and one on the Y chromosome. In line with previous research using other classes of genetic markers, our results indicate reduced Alu-associated insertion polymorphism on the human sex chromosomes, presumably reflective of the reduced recombination rates and lower effective population sizes on the sex chromosomes. The Alu insertion polymorphisms identified in this study should prove useful for the study of human population genetics.

We examine the distribution and structure of human genetic diversity for 710 individuals representing 31 populations from Africa, East Asia, Europe, and India using 100 Alu insertion polymorphisms from all 22 autosomes. Alu diversity is highest in Africans (0.349) and lowest in Europeans (0.297). Alu insertion frequency is lowest in Africans (0.463) and higher in Indians (0.544), E. Asians (0.557), and Europeans (0.559). Large genetic distances are observed among African populations and between African and non-African populations. The root of a neighbor-joining network is located closest to the African populations. These findings are consistent with an African origin of modern humans and with a bottleneck effect in the human populations that left Africa to colonize the rest of the world. Genetic distances among all pairs of populations show a significant product-moment correlation with geographic distances (r = 0.69, P < 0.00001). FST, the proportion of genetic diversity attributable to population subdivision is 0.141 for Africans/E. Asians/Europeans, 0.047 for E. Asians/Indians/Europeans, and 0.090 for all 31 populations. Resampling analyses show that ∼50 Alu polymorphisms are sufficient to obtain accurate and reliable genetic distance estimates. These analyses also demonstrate that markers with higher FST values have greater resolving power and produce more consistent genetic distance estimates.

A comprehensive analysis of two Alu Y lineage subfamilies was undertaken to assess Alu-associated genomic diversity and identify new Alu insertion polymorphisms for the study of human population genetics. Recently integrated Alu elements (283) from the Yg6 and Yi6 subfamilies were analyzed by polymerase chain reaction (PCR), and 25 of the loci analyzed were polymorphic for insertion presence/absence within the genomes of a diverse array of human populations. These newly identified Alu insertion polymorphisms will be useful tools for the study of human genomic diversity. Our screening of the Alu insertion loci also resulted in the recovery of several "young" Alu elements that resided at orthologous positions in nonhuman primate genomes. Sequence analysis demonstrated these "young" Alu insertions were the products of gene conversion events of older, preexisting Alu elements or independent parallel forward insertions of older Alu elements in the same short genomic region. The level of gene conversion between Alu elements suggests that it may have an influence on the single nucleotide polymorphism within Alu elements in the genome. We have also identified two genomic deletions associated with the retroposition and insertion of Alu Y lineage elements into the human genome. This type of Alu retroposition-mediated genomic deletion is a novel source of lineage-specific evolution within primate genomes.

CCR5 encodes a cell surface chemokine receptor molecule that serves as the principal coreceptor, with CD4, for HIV-type 1 (HIV-1). Varied HIV-1 susceptibility and time to progression to AIDS have been associated with polymorphisms in CCR5 . Many of these polymorphisms are located in the 5′ cis -regulatory region of CCR5 , suggesting that it may have been a target of natural selection. We characterized CCR5 sequence variation in this region in 400 chromosomes from worldwide populations and compared it to a genome-wide analysis of 100 Alu polymorphisms typed in the same populations. Variation was substantially higher than expected and characterized by an excess of intermediate-frequency alleles. A genealogy of CCR5 haplotypes had deep branch lengths despite markedly little differentiation among populations. This finding suggested a deviation from neutrality not accounted for by population structure, which was confirmed by tests for natural selection. These results are strong evidence that balancing selection has shaped the pattern of variation in CCR5 and suggest that HIV-1 resistance afforded by CCR5 5′ cis -regulatory region haplotypes may be the consequence of adaptive changes to older pathogens.

Variation in human skin/hair pigmentation is due to varied amounts of eumelanin (brown/black melanins) and phaeomelanin (red/yellow melanins) produced by the melanocytes. The melanocortin 1 receptor (MC1R) is a regulator of eu- and phaeomelanin production in the melanocytes, and MC1R mutations causing coat color changes are known in many mammals. We have sequenced the MC1R gene in 121 individuals sampled from world populations with an emphasis on Asian populations. We found variation at five nonsynonymous sites (resulting in the variants Arg67Gln, Asp84Glu, Val92Met, Arg151Cys, and Arg163Gln), but at only one synonymous site (A942G). Interestingly, the human consensus protein sequence is observed in all 25 African individuals studied, but at lower frequencies in the other populations examined, especially in East and Southeast Asians. The Arg163Gln variant is absent in the Africans studied, almost absent in Europeans, and at a low frequency (7%) in Indians, but is at an exceptionally high frequency (70%) in East and Southeast Asians. The MC1R gene in common and pygmy chimpanzees, gorilla, orangutan, and baboon was sequenced to study the evolution of MC1R. The ancestral human MC1R sequence is identical to the human consensus protein sequence, while MC1R varies considerably among higher primates. A comparison of the rates of substitution in genes in the melanocortin receptor family indicates that MC1R has evolved the fastest. In addition, the nucleotide diversity at the MC1R locus is shown to be several times higher than the average nucleotide diversity in human populations, possibly due to diversifying selection.

To examine the signature of population expansion on genetic variability at microsatellite loci, we consider a population that evolves according to the time-continuous Moran model, with growing population size and mutations that follow a general asymmetric stepwise mutation model. We present calculations of expected allele-size variance and homozygosity at a locus in such a model for several variants of growth, including stepwise, exponential, and logistic growth. These calculations in particular prove that population bottleneck followed by growth in size causes an imbalance between allele size variance and heterozygosity, characterized by the variance being transiently higher than expected under equilibrium conditions. This effect is, in a sense, analogous to that demonstrated before for the infinite allele model, where the number of alleles transiently increases after a stepwise growth of population. We analyze a set of data on tetranucleotide repeats that reveals the imbalance expected under the assumption of bottleneck followed by population growth in two out of three major racial groups. The imbalance is strongest in Asians, intermediate in Europeans, and absent in Africans. This finding is consistent with previous findings by others concerning the population expansion of modern humans, with the bottleneck event being most ancient in Africans, most recent in Asians, and intermediate in Europeans. Nevertheless, the imbalance index alone cannot reliably estimate the time of initiation of population expansion.

We have examined differences in diversity at 60 microsatellite loci among human population samples from three major continental groups to evaluate the hypothesis of greater African diversity in this rapidly evolving class of loci. Application of a statistical test that assumes equal mutation rates at all loci fails to demonstrate differences in microsatellite diversity, while a randomization test that does not make this assumption finds that Africans have significantly greater microsatellite diversity ( P < 10 −8 ) than do Asians and Europeans. Greater African diversity is most apparent at loci with smaller overall variance in allele size, suggesting that the record of population history has been erased at repeat loci with higher mutation rates. A power analysis shows that only 35–40 microsatellites are needed to establish this difference statistically, demonstrating the considerable evolutionary information contained in these systems. On average, African populations have ≈20% greater microsatellite diversity than do Asian and European populations. A comparison of continental diversity differences in microsatellites and mtDNA sequences suggests earlier demographic expansion of the ancestors of Africans.

Study Rationale: Unlike other cancer models, the risk factors predisposing patients to the development of Ewing's sarcoma and predicting subsequent disease behavior are largely unknown. Interestingly, the incidence of Ewing's sarcoma is highest in populations of European descent, which is 10X higher than the incidence observed in those of African descent. Furthermore, cure rates and overall survival are significantly worse in patients of African descent. Recently, it has been shown that the EWS/FLI fusion product selectively binds to a (GGAA)n microsatellite response element embedded within the promoter region of key EWS/FLI-target oncogenes. An increasing number of these repeats also appears to positively affect DNA binding and gene activation. We hypothesize that differences in polymorphic GGAA microsatellites within the promoter regions of key EWS/FLI-target genes may exist in African and European populations, contributing to observed differences in Ewing's sarcoma incidence and outcomes. Methods: Genomic DNA samples were obtained from 100 subjects each of African and European descent. The microsatellite regions within the promoter elements of two key EWS/FLI target genes (NR0B1 and CAV1) were PCR amplified, cloned and sequenced. Microsatellite sequences were analyzed and the overall length of the microsatellite, the total number of GGAA repeats and the maximal number of consecutive repeats were quantified. Data was statistically interpreted using a comparative two-tailed t-test. Results: Measurements of the CAV1 microsatellite response element were quite similar for both African and European subjects. In sharp contrast, marked inter-ethnic variation was observed for the NR0B1 GGAA microsatellite response element. African samples had a significantly longer GGAA microsatellite, a greater number of total (GGAA)n repeats and longer consecutive (GGAA)n repeats (p Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3979. doi:1538-7445.AM2012-3979