Lynn B. Jorde

Analysis of Genetic Inheritance in a Family Quartet by Whole-Genome Sequencing

Runs in the Family The power to detect mutations involved in disease by genome sequencing is enhanced when combined with the ability to discover specific mutations that may have arisen between offspring and parents. Roach et al. (p. 636, published online 10 March) present the sequence of a family with two offspring affected with two genetic disorders: Miller syndrome and primary ciliary dyskinesia. Sequence analysis of the children and their parents not only showed that the intergenerational mutation rate was lower than anticipated but also revealed recombination sites and the occurrence of rare polymorphisms. Genomic sequencing of an entire family reveals the rate of spontaneous mutations in humans and identifies disease genes. We analyzed the whole-genome sequences of a family of four, consisting of two siblings and their parents. Family-based sequencing allowed us to delineate recombination sites precisely, identify 70% of the sequencing errors (resulting in > 99.999% accuracy), and identify very rare single-nucleotide polymorphisms. We also directly estimated a human intergeneration mutation rate of ~1.1 × 10−8 per position per haploid genome. Both offspring in this family have two recessive disorders: Miller syndrome, for which the gene was concurrently identified, and primary ciliary dyskinesia, for which causative genes have been previously identified. Family-based genome analysis enabled us to narrow the candidate genes for both of these Mendelian disorders to only four. Our results demonstrate the value of complete genome sequencing in families.

Genetic Evidence for High-Altitude Adaptation in Tibet

No Genetic Vertigo Peoples living in high altitudes have adapted to their situation (see the Perspective by Storz). To identify gene regions that might have contributed to high-altitude adaptation in Tibetans, Simonson et al. (p. 72, published online 13 May) conducted a genome scan of nucleotide polymorphism comparing Tibetans, Han Chinese, and Japanese, while Yi et al. (p. 75) performed comparable analyses on the coding regions of all genes—their exomes. Both studies converged on a gene, endothelial Per-Arnt-Sim domain protein 1 (also known as hypoxia-inducible factor 2α), which has been linked to the regulation of red blood cell production. Other genes identified that were potentially under selection included adult and fetal hemoglobin and two functional candidate loci that were correlated with low hemoglobin concentration in Tibetans. Future detailed functional studies will now be required to examine the mechanistic underpinnings of physiological adaptation to high altitudes. A candidate gene approach reveals genes under selection in humans living at high altitudes. Tibetans have lived at very high altitudes for thousands of years, and they have a distinctive suite of physiological traits that enable them to tolerate environmental hypoxia. These phenotypes are clearly the result of adaptation to this environment, but their genetic basis remains unknown. We report genome-wide scans that reveal positive selection in several regions that contain genes whose products are likely involved in high-altitude adaptation. Positively selected haplotypes of EGLN1 and PPARA were significantly associated with the decreased hemoglobin phenotype that is unique to this highland population. Identification of these genes provides support for previously hypothesized mechanisms of high-altitude adaptation and illuminates the complexity of hypoxia-response pathways in humans.

The Distribution of Human Genetic Diversity: A Comparison of Mitochondrial, Autosomal, and Y-Chromosome Data

We report a comparison of worldwide genetic variation among 255 individuals by using autosomal, mitochondrial, and Y-chromosome polymorphisms. Variation is assessed by use of 30 autosomal restriction-site polymorphisms (RSPs), 60 autosomal short-tandem-repeat polymorphisms (STRPs), 13 Alu-insertion polymorphisms and one LINE-1 element, 611 bp of mitochondrial control-region sequence, and 10 Y-chromosome polymorphisms. Analysis of these data reveals substantial congruity among this diverse array of genetic systems. With the exception of the autosomal RSPs, in which an ascertainment bias exists, all systems show greater gene diversity in Africans than in either Europeans or Asians. Africans also have the largest total number of alleles, as well as the largest number of unique alleles, for most systems. GST values are 11%-18% for the autosomal systems and are two to three times higher for the mtDNA sequence and Y-chromosome RSPs. This difference is expected because of the lower effective population size of mtDNA and Y chromosomes. A lower value is seen for Y-chromosome STRs, reflecting a relative lack of continental population structure, as a result of rapid mutation and genetic drift. Africa has higher GST values than does either Europe or Asia for all systems except the Y-chromosome STRs and Alus. All systems except the Y-chromosome STRs show less variation between populations within continents than between continents. These results are reassuring in their consistency and offer broad support for an African origin of modern human populations.

Human Population Genetic Structure and Inference of Group Membership

A major goal of biomedical research is to develop the capability to provide highly personalized health care. To do so, it is necessary to understand the distribution of interindividual genetic variation at loci underlying physical characteristics, disease susceptibility, and response to treatment. Variation at these loci commonly exhibits geographic structuring and may contribute to phenotypic differences between groups. Thus, in some situations, it may be important to consider these groups separately. Membership in these groups is commonly inferred by use of a proxy such as place-of-origin or ethnic affiliation. These inferences are frequently weakened, however, by use of surrogates, such as skin color, for these proxies, the distribution of which bears little resemblance to the distribution of neutral genetic variation. Consequently, it has become increasingly controversial whether proxies are sufficient and accurate representations of groups inferred from neutral genetic variation. This raises three questions: how many data are required to identify population structure at a meaningful level of resolution, to what level can population structure be resolved, and do some proxies represent population structure accurately? We assayed 100 Alu insertion polymorphisms in a heterogeneous collection of approximately 565 individuals, approximately 200 of whom were also typed for 60 microsatellites. Stripped of identifying information, correct assignment to the continent of origin (Africa, Asia, or Europe) with a mean accuracy of at least 90% required a minimum of 60 Alu markers or microsatellites and reached 99%-100% when >/=100 loci were used. Less accurate assignment (87%) to the appropriate genetic cluster was possible for a historically admixed sample from southern India. These results set a minimum for the number of markers that must be tested to make strong inferences about detecting population structure among Old World populations under ideal experimental conditions. We note that, whereas some proxies correspond crudely, if at all, to population structure, the heuristic value of others is much higher. This suggests that a more flexible framework is needed for making inferences about population structure and the utility of proxies.

Clinical and Biochemical Abnormalities in People Heterozygous for Hemochromatosis

BACKGROUND Ten percent of whites are heterozygous for the HLA-linked hemochromatosis mutation. We performed a cross-sectional analysis of 1058 genotyped heterozygotes to define the effects of age and sex on the phenotype. METHODS The heterozygous genotype was assigned to 505 male and 553 female members of 202 pedigrees, each with an HLA-typed homozygous proband. We measured serum iron, transferrin saturation, and ferritin in all heterozygotes and in 321 genetically normal subjects (unaffected family members or spouses of family members). Liver biopsies were performed in a subgroup of heterozygotes. RESULTS The mean serum iron concentrations and transferrin-saturation values were higher in heterozygotes than in normal subjects and did not increase with age. Initial transferrin-saturation levels exceeding the threshold associated with the homozygous genotype were found in 4 percent of male and 8 percent of female heterozygotes. The geometric mean serum ferritin concentration was higher in heterozygotes than in normal subjects and increased with age. Higher-than-normal values were found in 20 percent of male and 8 percent of female heterozygotes. The clinical and biochemical expression of hemochromatosis was more marked in heterozygotes with paternally transmitted mutations than in those with maternally transmitted mutations. Liver-biopsy abnormalities were generally associated with alcohol abuse, hepatitis, or porphyria cutanea tarda. CONCLUSIONS The phenotype of persons heterozygous for hemochromatosis differs from that of normal subjects, but complications due to iron overload alone in these heterozygotes are extremely rare.

De novo mutations in ATP1A3 cause alternating hemiplegia of childhood

Alternating hemiplegia of childhood (AHC) is a rare, severe neurodevelopmental syndrome characterized by recurrent hemiplegic episodes and distinct neurological manifestations. AHC is usually a sporadic disorder and has unknown etiology. We used exome sequencing of seven patients with AHC and their unaffected parents to identify de novo nonsynonymous mutations in ATP1A3 in all seven individuals. In a subsequent sequence analysis of ATP1A3 in 98 other patients with AHC, we found that ATP1A3 mutations were likely to be responsible for at least 74% of the cases; we also identified one inherited mutation in a case of familial AHC. Notably, most AHC cases are caused by one of seven recurrent ATP1A3 mutations, one of which was observed in 36 patients. Unlike ATP1A3 mutations that cause rapid-onset dystonia-parkinsonism, AHC-causing mutations in this gene caused consistent reductions in ATPase activity without affecting the level of protein expression. This work identifies de novo ATP1A3 mutations as the primary cause of AHC and offers insight into disease pathophysiology by expanding the spectrum of phenotypes associated with mutations in ATP1A3.

Mobile elements create structural variation: Analysis of a complete human genome

Structural variants (SVs) are common in the human genome. Because approximately half of the human genome consists of repetitive, transposable DNA sequences, it is plausible that these elements play an important role in generating SVs in humans. Sequencing of the diploid genome of one individual human (HuRef) affords us the opportunity to assess, for the first time, the impact of mobile elements on SVs in an individual in a thorough and unbiased fashion. In this study, we systematically evaluated more than 8000 SVs to identify mobile element-associated SVs as small as 100 bp and specific to the HuRef genome. Combining computational and experimental analyses, we identified and validated 706 mobile element insertion events (including Alu, L1, SVA elements, and nonclassical insertions), which added more than 305 kb of new DNA sequence to the HuRef genome compared with the Human Genome Project (HGP) reference sequence (hg18). We also identified 140 mobile element-associated deletions, which removed approximately 126 kb of sequence from the HuRef genome. Overall, approximately 10% of the HuRef-specific indels larger than 100 bp are caused by mobile element-associated events. More than one-third of the insertion/deletion events occurred in genic regions, and new Alu insertions occurred in exons of three human genes. Based on the number of insertions and the estimated time to the most recent common ancestor of HuRef and the HGP reference genome, we estimated the Alu, L1, and SVA retrotransposition rates to be one in 21 births, 212 births, and 916 births, respectively. This study presents the first comprehensive analysis of mobile element-related structural variants in the complete DNA sequence of an individual and demonstrates that mobile elements play an important role in generating inter-individual structural variation.

Disease-Related Conditions in Relatives of Patients with Hemochromatosis

BACKGROUND Hemochromatosis occurs in approximately 5 white people per 1000 and is usually due to homozygosity for mutations in the HLA-linked HFE gene. Although screening has been proposed, the proportion of homozygotes with conditions related to hemochromatosis is uncertain. METHODS We studied the prevalence of disease-related conditions among relatives of probands with hemochromatosis. We identified probands who presented to a clinic with signs or symptoms of hemochromatosis or who had elevated transferrin-saturation values. We identified homozygous relatives, mainly siblings, on the basis of HLA identity with the proband and by HFE genotyping. Disease-related conditions were cirrhosis, hepatic fibrosis, elevated amino-transferase values, and hemochromatotic arthropathy. RESULTS We identified 214 homozygous relatives of 291 homozygous probands. Of the 113 men in this group (mean age, 41 years), 96 (85 percent) had iron overload, and 43 (38 percent) had at least one disease-related condition. Of the 52 men over 40 years of age, 27 (52 percent) had at least one disease-related condition. Of the 101 female homozygous relatives (mean age, 44 years), 69 (68 percent) had iron overload, and 10 (10 percent) had at least one disease-related condition. Of the 43 women over 50 years of age, 7 (16 percent) had at least one disease-related condition. If the proband had a disease-related condition, relatives who were men were more likely to have morbidity than if the proband had no disease-related condition. CONCLUSIONS A substantial number of homozygous relatives of patients with hemochromatosis--more commonly men than women--have conditions related to hemochromatosis that have yet to be detected clinically.

The Simons Genome Diversity Project: 300 genomes from 142 diverse populations

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.

A Comprehensive Map of Mobile Element Insertion Polymorphisms in Humans

As a consequence of the accumulation of insertion events over evolutionary time, mobile elements now comprise nearly half of the human genome. The Alu, L1, and SVA mobile element families are still duplicating, generating variation between individual genomes. Mobile element insertions (MEI) have been identified as causes for genetic diseases, including hemophilia, neurofibromatosis, and various cancers. Here we present a comprehensive map of 7,380 MEI polymorphisms from the 1000 Genomes Project whole-genome sequencing data of 185 samples in three major populations detected with two detection methods. This catalog enables us to systematically study mutation rates, population segregation, genomic distribution, and functional properties of MEI polymorphisms and to compare MEI to SNP variation from the same individuals. Population allele frequencies of MEI and SNPs are described, broadly, by the same neutral ancestral processes despite vastly different mutation mechanisms and rates, except in coding regions where MEI are virtually absent, presumably due to strong negative selection. A direct comparison of MEI and SNP diversity levels suggests a differential mobile element insertion rate among populations.