ALFRED: an allele frequency database for diverse populations and DNA polymorphisms—an update

ALFRED (the ALelle FREquency Database) is designed to store and disseminate frequencies of alleles at human polymorphic sites for multiple populations, primarily for the population genetics and molecular anthropology communities. Currently ALFRED has information on over 180 polymorphic sites for more than 70 populations. Since our initial release of the database we have focussed on increasing the quantity and quality of data, making reciprocal links between ALFRED and other related databases, and providing useful tools to make the data more comprehensible to the end user. ALFRED is accessible from the Kidd Lab home page (http://info.med.yale.edu/genetics/kkidd/) or from ALFRED directly (http://alfred.med.yale.edu/alfred/index.asp).

Social transmission of reproductive behavior increases frequency of inherited disorders in a young-expanding population

The observation of high frequencies of certain inherited disorders in the population of Saguenay–Lac Saint Jean can be explained in terms of the variance and the correlation of effective family size (EFS) from one generation to the next. We have shown this effect by using the branching process approach with real demographic data. When variance of EFS is included in the model, despite its profound effect on mutant allele frequency, any mutant introduced in the population never reaches the known carrier frequencies (between 0.035 and 0.05). It is only when the EFS correlation between generations is introduced into the model that we can explain the rise of the mutant alleles. This correlation is described by a c parameter that reflects the dependency of children’s EFS on their parents’ EFS. The c parameter can be considered to reflect social transmission of demographic behavior. We show that such social transmission dramatically reduces the effective population size. This could explain particular distributions in allele frequencies and unusually high frequency of certain inherited disorders in some human populations.

Frequency of resistance to Bacillus thuringiensis in field populations of pink bollworm

Strategies for delaying pest resistance to genetically modified crops that produce Bacillus thuringiensis (Bt) toxins are based primarily on theoretical models. One key assumption of such models is that genes conferring resistance are rare. Previous estimates for lepidopteran pests targeted by Bt crops seem to meet this assumption. We report here that the estimated frequency of a recessive allele conferring resistance to Bt toxin Cry1Ac was 0.16 (95% confidence interval = 0.05–0.26) in strains of pink bollworm (Pectinophora gossypiella) derived from 10 Arizona cotton fields during 1997. Unexpectedly, the estimated resistance allele frequency did not increase from 1997 to 1999 and Bt cotton remained extremely effective against pink bollworm. These results demonstrate that the assumptions and predictions of resistance management models must be reexamined.

A pre-Columbian Y chromosome-specific transition and its implications for human evolutionary history.

A polymorphic C-->T transition located on the human Y chromosome was found by the systematic comparative sequencing of Y-specific sequence-tagged sites by denaturing high-performance liquid chromatography. The results of genotyping representative global indigenous populations indicate that the locus is polymorphic exclusively within the Western Hemisphere. The pre-Columbian T allele occurs at > 90% frequency within the native South and Central American populations examined, while its occurrence in North America is approximately 50%. Concomitant genotyping at the polymorphic tetranucleotide microsatellite DYS19 locus revealed that the C-->T mutation displayed significant linkage disequilibrium with the 186-bp allele. The data suggest a single origin of linguistically diverse native Americans with subsequent haplotype differentiation within radiating indigenous populations as well as post-Columbian European and African gene flow. The mutation may have originated either in North America at a very early time during the expansion or before it, in the ancestral population(s) from which all Americans may have originated. The analysis of linkage of the DYS199 and the DYS19 tetranucleotide loci suggests that the C-->T mutation may have occurred around 30,000 years ago. We estimate the nucleotide diversity over 4.2 kb of the nonrecombining portion of the Y chromosome to be 0.00014. compared to autosomes, the majority of variation is due to the smaller effective population size of the Y chromosome rather than selective sweeps. There begins to emerge a pattern of pronounced geographical localization of Y-specific nucleotide substitution polymorphisms.

Fetal origins of the TEL-AML1 fusion gene in identical twins with leukemia

The TEL (ETV6)−AML1 (CBFA2) gene fusion is the most common reciprocal chromosomal rearrangement in childhood cancer occurring in ≈25% of the most predominant subtype of leukemia— common acute lymphoblastic leukemia. The TEL-AML1 genomic sequence has been characterized in a pair of monozygotic twins diagnosed at ages 3 years, 6 months and 4 years, 10 months with common acute lymphoblastic leukemia. The twin leukemic DNA shared the same unique (or clonotypic) but nonconstitutive TEL-AML1 fusion sequence. The most plausible explanation for this finding is a single cell origin of the TEL-AML fusion in one fetus in utero, probably as a leukemia-initiating mutation, followed by intraplacental metastasis of clonal progeny to the other twin. Clonal identity is further supported by the finding that the leukemic cells in the two twins shared an identical rearranged IGH allele. These data have implications for the etiology and natural history of childhood leukemia.

Gilbert syndrome and glucose-6-phosphate dehydrogenase deficiency: A dose-dependent genetic interaction crucial to neonatal hyperbilirubinemia

Severe jaundice leading to kernicterus or death in the newborn is the most devastating consequence of glucose-6-phosphate dehydrogenase (EC 1.1.1.49; G-6-PD) deficiency. We asked whether the TA repeat promoter polymorphism in the gene for uridinediphosphoglucuronate glucuronosyltransferase 1 (EC 2.4.1.17; UDPGT1), associated with benign jaundice in adults (Gilbert syndrome), increases the incidence of neonatal hyperbilirubinemia in G-6-PD deficiency. DNA from term neonates was analyzed for UDPGT1 polymorphism (normal homozygotes, heterozygotes, variant homozygotes), and for G-6-PD Mediterranean deficiency. The variant UDPGT1 promoter allele frequency was similar in G-6-PD-deficient and normal neonates. Thirty (22.9%) G-6-PD deficient neonates developed serum total bilirubin ≥ 257 μmol/liter, vs. 22 (9.2%) normals (P = 0.0005). Of those with the normal homozygous UDPGT1 genotype, the incidence of hyperbilirubinemia was similar in G-6-PD-deficients and controls (9.7% and 9.9%). In contrast, in the G-6-PD-deficient neonates, those with the heterozygous or homozygous variant UDPGT1 genotype had a higher incidence of hyperbilirubinemia than corresponding controls (heterozygotes: 31.6% vs. 6.7%, P < 0.0001; variant homozygotes: 50% vs. 14.7%, P = 0.02). Among G-6-PD-deficient infants the incidence of hyperbilirubinemia was greater in those with the heterozygous (31.6%, P = 0.006) or variant homozygous (50%, P = 0.003) UDPGT1 genotype than in normal homozygotes. In contrast, among those normal for G-6-PD, the UDPGT1 polymorphism had no significant effect (heterozygotes: 6.7%; variant homozygotes: 14.7%). Thus, neither G-6-PD deficiency nor the variant UDPGT1 promoter, alone, increased the incidence of hyperbilirubinemia, but both in combination did. This gene interaction may serve as a paradigm of the interaction of benign genetic polymorphisms in the causation of disease.

Emergence of FY*Anull in a Plasmodium vivax-endemic region of Papua New Guinea

In Papua New Guinea (PNG), numerous blood group polymorphisms and hemoglobinopathies characterize the human population. Human genetic polymorphisms of this nature are common in malarious regions, and all four human malaria parasites are holoendemic below 1500 meters in PNG. At this elevation, a prominent condition characterizing Melanesians is α+-thalassemia. Interestingly, recent epidemiological surveys have demonstrated that α+-thalassemia is associated with increased susceptibility to uncomplicated malaria among young children. It is further proposed that α+-thalassemia may facilitate so-called “benign” Plasmodium vivax infection to act later in life as a “natural vaccine” against severe Plasmodium falciparum malaria. Here, in a P. vivax-endemic region of PNG where the resident Abelam-speaking population is characterized by a frequency of α+-thalassemia ≥0.98, we have discovered the mutation responsible for erythrocyte Duffy antigen-negativity (Fy[a−b−]) on the FY*A allele. In this study population there were 23 heterozygous and no homozygous individuals bearing this new allele (allele frequency, 23/1062 = 0.022). Flow cytometric analysis illustrated a 2-fold difference in erythroid-specific Fy-antigen expression between heterozygous (FY*A/FY*Anull) and homozygous (FY*A/FY*A) individuals, suggesting a gene-dosage effect. In further comparisons, we observed a higher prevalence of P. vivax infection in FY*A/FY*A (83/508 = 0.163) compared with FY*A/FY*Anull (2/23 = 0.087) individuals (odds ratio = 2.05, 95% confidence interval = 0.47–8.91). Emergence of FY*Anull in this population suggests that P. vivax is involved in selection of this erythroid polymorphism. This mutation would ultimately compromise α+-thalassemia/P. vivax-mediated protection against severe P. falciparum malaria.

Functional and physiological consequences of genetic variation at phosphoglucose isomerase: Heat shock protein expression is related to enzyme genotype in a montane beetle

Allele frequency variation at the phosphoglucose isomerase (PGI) locus in Californian populations of the beetle Chrysomela aeneicollis suggests that PGI may be undergoing natural selection. We quantified (i) apparent Michaelis-Menten constant (Km) of fructose 6-phosphate at different temperatures and (ii) thermal stability for three common PGI genotypes (1–1, 1–4, and 4–4). We also measured air temperature (Ta) and beetle body temperature (Tb) in three montane drainages in the Sierra Nevada, California. Finally, we measured 70-kDa heat shock protein (Hsp70) expression in field-collected and laboratory-acclimated beetles. We found that PGI allele 1 predominated in the northernmost drainage, Rock Creek (RC), which was also significantly cooler than the southernmost drainage, Big Pine Creek (BPC), where PGI allele 4 predominated. Allele frequencies and air temperatures were intermediate in the middle drainage, Bishop Creek (BC). Differences among genotypes in Km (1–1 > 1–4 > 4–4) and thermal stability (4–4 > 1–4 > 1–1) followed a pattern consistent with temperature adaptation. In nature, Tb was closely related to Ta. Hsp70 expression in adult beetles decreased with elevation and differed among drainages (BPC > BC > RC). After laboratory acclimation (8 days, 20°C day, 4°C night) and heat shock (4 h, 28–36°C), Hsp70 expression was greater for RC than BPC beetles. In RC, field-collected beetles homozygous for PGI 1–1 had higher Hsp70 levels than heterozygotes or a 4–4 homozygote. These results reveal functional and physiological differences among PGI genotypes, which suggest that montane populations of this beetle are locally adapted to temperature.

Neutral behavior of shared polymorphism

Several cases have been described in the literature where genetic polymorphism appears to be shared between a pair of species. Here we examine the distribution of times to random loss of shared polymorphism in the context of the neutral Wright–Fisher model. Order statistics are used to obtain the distribution of times to loss of a shared polymorphism based on Kimura’s solution to the diffusion approximation of the Wright–Fisher model. In a single species, the expected absorption time for a neutral allele having an initial allele frequency of ½ is 2.77 N generations. If two species initially share a polymorphism, that shared polymorphism is lost as soon as either of two species undergoes fixation. The loss of a shared polymorphism thus occurs sooner than loss of polymorphism in a single species and has an expected time of 1.7 N generations. Molecular sequences of genes with shared polymorphism may be characterized by the count of the number of sites that segregate in both species for the same nucleotides (or amino acids). The distribution of the expected numbers of these shared polymorphic sites also is obtained. Shared polymorphism appears to be more likely at genetic loci that have an unusually large number of segregating alleles, and the neutral coalescent proves to be very useful in determining the probability of shared allelic lineages expected by chance. These results are related to examples of shared polymorphism in the literature.

A rat genetic map constructed by representational difference analysis markers with suitability for large-scale typing.

Representational difference analysis (RDA) was applied to isolate chromosomal markers in the rat. Four series of RDA [restriction enzymes, BamHI and HindIII; subtraction of ACI/N (ACI) amplicon from BUF/Nac (BUF) amplicon and vice versa] yielded 131 polymorphic markers; 125 of these markers were mapped to all chromosomes except for chromosome X. This was done by using a mapping panel of 105 ACI x BUF F2 rats. To complement the relative paucity of chromosomal markers in the rat, genetically directed RDA, which allows isolation of polymorphic markers in the specific chromosomal region, was performed. By changing the F2 driver-DNA allele frequency around the region, four markers were isolated from the D1Ncc1 locus. Twenty-five of 27 RDA markers were informative regarding the dot blot analysis of amplicons, hybridizing only with tester amplicons. Dot blot analysis at a high density per unit of area made it possible to process a large number of samples. Quantitative trait loci can now be mapped in the rat genome by processing a large number of samples with RDA markers and then by isolating markers close to the loci of interest by genetically directed RDA.

Frequency of HLA allele-specific peptide motifs in HIV-1 proteins correlates with the allele’s association with relative rates of disease progression after HIV-1 infection

An HLA allele-specific cytotoxic T lymphocyte response is thought to influence the rate of disease progression in HIV-1-infected individuals. In a prior study of 139 HIV-1-infected homosexual men, we identified HLA class I alleles and observed an association of specific alleles with different relative hazards for progression to AIDS. Seeking an explanation for this association, we searched HIV-1 protein sequences to determine the number of peptides matching motifs defined by combinations of specific amino acids reported to bind 16 class I alleles. Analyzing complete sequences of 12 clade B HIV isolates, we determined the number of allele motifs that were conserved (occurring in all 12 isolates) and nonconserved (occurring in only one isolate), as well as the average number of allele motifs per isolate. We found significant correlations with an allele’s association with disease progression for counts of conserved motifs in gag (R = 0.73; P = 0.002), pol (R = 0.58, P = 0.024), gp120 (R = 0.78, P = 0.00056), and total viral protein sequences (R = 0.67, P = 0.0058) and also for counts of nonconserved motifs in gag (R = 0.62, P = 0.013), pol (R = 0.74, P = 0.0017), gp41 (R = 0.52, P = 0.046), and total viral protein (R = 0.71, P = 0.0033). We also found significant correlations for the average number of motifs per isolate for gag, pol, gp120, and total viral protein. This study provides a plausible functional explanation for the observed association of different HLA alleles with variable rates of disease progression.