X-linked Loci Research Articles

Genomic Evidence for Island Population Conversion Resolves Conflicting Theories of Polar Bear Evolution

Despite extensive genetic analysis, the evolutionary relationship between polar bears (Ursus maritimus) and brown bears (U. arctos) remains unclear. The two most recent comprehensive reports indicate a recent divergence with little subsequent admixture or a much more ancient divergence followed by extensive admixture. At the center of this controversy are the Alaskan ABC Islands brown bears that show evidence of shared ancestry with polar bears. We present an analysis of genome-wide sequence data for seven polar bears, one ABC Islands brown bear, one mainland Alaskan brown bear, and a black bear (U. americanus), plus recently published datasets from other bears. Surprisingly, we find clear evidence for gene flow from polar bears into ABC Islands brown bears but no evidence of gene flow from brown bears into polar bears. Importantly, while polar bears contributed <1% of the autosomal genome of the ABC Islands brown bear, they contributed 6.5% of the X chromosome. The magnitude of sex-biased polar bear ancestry and the clear direction of gene flow suggest a model wherein the enigmatic ABC Island brown bears are the descendants of a polar bear population that was gradually converted into brown bears via male-dominated brown bear admixture. We present a model that reconciles heretofore conflicting genetic observations. We posit that the enigmatic ABC Islands brown bears derive from a population of polar bears likely stranded by the receding ice at the end of the last glacial period. Since then, male brown bear migration onto the island has gradually converted these bears into an admixed population whose phenotype and genotype are principally brown bear, except at mtDNA and X-linked loci. This process of genome erosion and conversion may be a common outcome when climate change or other forces cause a population to become isolated and then overrun by species with which it can hybridize.

Identification of a novel imprinting mechanism at the X-linked imprinted locus, X-linked Lymphocyte Regulated 3/4 (Xlr3/4)

Background Genomic imprinting is the epigenetic process by which a subset of genes is expressed from only one allele based on the parent of origin. To date there are approximately 150 confirmed autosomal imprinted loci in the mammalian genome. The epigenetic hallmark of an autosomal imprinted locus is the presence of differential methylation at CpG islands otherwise known as a DMR[1]. Imprint maintenance at autosomal loci is accomplished through specific histone modifications[2]. In 2005, Raefski et.al. found the first X-linked imprinted locus, Xlr3/4 of which three paralogs, Xlr3b/4b/4c, were maternally expressed and paternally silenced in mouse neonatal brain[3]. Here we use a series of techniques to uncover the imprinting mechanism at the Xlr locus using known autosomal imprinting mechanisms as a model.

English

&nbsp; Here we report a mammal sexing procedure based on the detection of quantitative differences between females and males in the X-linked loci (quantitative sexing, Q-sexing). This novel technique was validated using samples from Siberian tigers (Panthera tigris altaica) whose sexes were known. The Q-sexing technique relies on the fact that amplifications proceeding exclusively from the two X chromosomes in a female mammal should reach the threshold cycle (CT) in a real-time quantitative real time polymerase chain reaction (qPCR) assay sooner than amplifications from the single male X chromosome. Nevertheless, given that the amplification efficiency may vary between samples, results have to be calibrated to a marker that does not vary in copy number between the sexes (for example, an autosomal-linked locus). For this purpose we used quantitative real time polymerase chain reaction (RT qPCR) assays to quantify the amount of three specific Siberian tiger microsatellite markers (X-/Y- and autosomal-linked loci) in individual samples in order to determine the sex of an animal. A difference of one CT&nbsp;between the X and the autosome-linked loci was detected in males, but no such difference was present in female samples. The Q-sexing technique unambiguously separates female from male Siberian tigers. The future of RT qPCR is bright as technology is becoming ever more rapid, cost-effective, easier to use and capable of processing higher throughputs. Thus, we expect that our novel technique for animal sexing will have a wide applicability, although further studies are still needed to adapt it to other animal species using specific primers. &nbsp; Key words:&nbsp;Polymerase chain reaction (PCR), quantitative real time polymerase chain reaction (qPCR), quantitative sexing, Siberian tiger.

Genomic Imprinting Leads to Less Selectively Maintained Polymorphism on X Chromosomes

Population-genetic models are developed to investigate the consequences of viability selection at a diallelic X-linked locus subject to genomic imprinting. Under complete paternal-X inactivation, a stable polymorphism is possible under the same conditions as for paternal-autosome inactivation with differential selection on males and females. A necessary but not sufficient condition is that there is sexual conflict, with selection acting in opposite directions in males and females. In contrast, models of complete maternal-X inactivation never admit a stable polymorphism and alleles will either be fixed or lost from the population. Models of complete paternal-X inactivation are more complex than corresponding models of maternal-X inactivation, as inactivation of paternally derived X chromosomes in females screens these chromosomes from selection for a generation. We also demonstrate that polymorphism is possible for incomplete X inactivation, but that the parameter conditions are more restrictive than for complete paternal-X inactivation. Finally, we investigate the effects of recurrent mutation in our models and show that deleterious alleles in mutation-selection balance at imprinted X-linked loci are at frequencies rather similar to those with corresponding selection pressures and mutation rates at unimprinted loci. Overall, our results add to the reasons for expecting less selectively maintained allelic variation on X chromosomes.

X chromosome inactivation pattern in BRCA gene mutation carriers

An association of preferential X chromosome inactivation (XCI) with BRCA gene status and breast/ovarian cancer risk has been reported. We evaluated XCI in a large group of BRCA mutation carriers compared to non-carriers and investigated associations between preferential XCI (⩾90:10) and age, mutated gene, cancer development and chemotherapy. XCI was analysed by human androgen receptor (HUMARA) assay and pyrosequencing in 437 BRCA1 or BRCA2 mutation carriers and 445 age-matched controls. The distribution of XCI patterns in the two groups was compared by logistic regression analysis. The association between preferential XCI and selected variables was investigated in both univariate and multivariate fashion. In univariate analyses preferential XCI was not significantly associated with the probability of being a BRCA mutation carrier, nor with cancer status, whereas chemotherapeutic regime and age both showed a significant association. In multivariate analysis only age maintained significance (odds ratio, 1.056; 95% confidence interval, 1.016–1.096). Our findings do not support the usefulness of XCI analysis for the identification of BRCA mutation carriers and cancer risk assessment. The increasing preferential XCI frequency with ageing and the association with chemotherapy justify extending the investigation to other categories of female cancer patients to identify possible X-linked loci implicated in cell survival.

The mouse hybrid zone in Central Europe: from morphology to molecules

Despite the long-term study of the house mouse hybrid zone in Europe knowledge of its course in some areas is still rather vague. Comparisons of different portions of the zone showed some common patterns, however, several discordances were also revealed, the most remarkable being introgression of the Y chromosome. We sampled mice along the presumed course of the secondary contact zone between two subspecies, Mus musculus musculus and M. m. domesticus, from Schleswig-Holstein to southern Bavaria, in order to localize more precisely its position. A second aim was to reveal whether introgression shows some general rules obscured until now by studies of geographically isolated transects of the zone. We employed maternally (mtDNA), paternally (Y), and biparentally inherited markers and related their introgression patterns to the hybrid index (HI) based on five X-linked loci. While transition of autosomal loci across the zone was congruent with changes in HI, mtDNA showed bidirectional introgression with alien alleles occurring far behind the zone. Finally, the Y chromosome displayed asymmetric unidirectional introgression of the musculus type into domesticus background. We discuss evolutionary forces shaping these patterns.

Faster-X evolution of gene expression in Drosophila.

DNA sequences on X chromosomes often have a faster rate of evolution when compared to similar loci on the autosomes, and well articulated models provide reasons why the X-linked mode of inheritance may be responsible for the faster evolution of X-linked genes. We analyzed microarray and RNA–seq data collected from females and males of six Drosophila species and found that the expression levels of X-linked genes also diverge faster than autosomal gene expression, similar to the “faster-X” effect often observed in DNA sequence evolution. Faster-X evolution of gene expression was recently described in mammals, but it was limited to the evolutionary lineages shortly following the creation of the therian X chromosome. In contrast, we detect a faster-X effect along both deep lineages and those on the tips of the Drosophila phylogeny. In Drosophila males, the dosage compensation complex (DCC) binds the X chromosome, creating a unique chromatin environment that promotes the hyper-expression of X-linked genes. We find that DCC binding, chromatin environment, and breadth of expression are all predictive of the rate of gene expression evolution. In addition, estimates of the intraspecific genetic polymorphism underlying gene expression variation suggest that X-linked expression levels are not under relaxed selective constraints. We therefore hypothesize that the faster-X evolution of gene expression is the result of the adaptive fixation of beneficial mutations at X-linked loci that change expression level in cis. This adaptive faster-X evolution of gene expression is limited to genes that are narrowly expressed in a single tissue, suggesting that relaxed pleiotropic constraints permit a faster response to selection. Finally, we present a conceptional framework to explain faster-X expression evolution, and we use this framework to examine differences in the faster-X effect between Drosophila and mammals.

A major X-linked locus affects kidney function in mice

Chronic kidney disease is a common disease with increasing prevalence in the western population. One common reason for chronic kidney failure is diabetic nephropathy. Diabetic nephropathy and hyperglycemia are characteristics of the mouse inbred strain KK/HlJ, which is predominantly used as a model for metabolic syndrome due to its inherited glucose intolerance and insulin resistance. We used KK/HlJ, an albuminuria-sensitive strain, and C57BL/6J, an albuminuria-resistant strain, to perform a quantitative trait locus (QTL) cross to identify the genetic basis for chronic kidney failure. Albumin-creatinine ratio (ACR) was measured in 130 F2 male offspring. One significant QTL was identified on chromosome (Chr) X and four suggestive QTL were found on Chrs 6, 7, 12, and 13. Narrowing of the QTL region was focused on the X-linked QTL and performed by incorporating genotype and expression analyses for genes located in the region. From the 485 genes identified in the X-linked QTL region, a few candidate genes were identified using a combination of bioinformatic evidence based on genomic comparison of the parental strains and known function in urine homeostasis. Finally, this study demonstrates the significance of the X chromosome in the genetic determination of albuminuria.

Gene Expression and DNA Methylation Status of Chicken Primordial Germ Cells

DNA methylation reprogramming of primordial germ cells (PGCs) in mammals establishes monoallelic expression of imprinting genes, maintains retrotransposons in an inactive state, inactivates one of the two X chromosomes, and suppresses gene expression. However, the roles of DNA methylation in chickens PGCs are unknown. In this study, we found a 1.5-fold or greater difference in the expression of 261 transcripts when comparing PGCs and chicken embryonic fibroblasts (CEFs) using an Affymetrix GeneChip Chicken Genome Array. In addition, we analyzed the methylation patterns of the regions ~5-kb upstream of 261 sorted genes, 51 of which were imprinting homologous loci and 49 of which were X-linked homologous loci in chicken using the MeDIP Array by Roche NimbleGen. Seven hypomethylated and five hypermethylated regions within the 5-kb upstream regions of 261 genes were found in PGCs when compared with CEFs. These differentially methylated regions were restrictively matched to differentially expressed genes in PGCs. We also detected 203 differentially methylated regions within imprinting and X-linked homologous regions between male PGCs and female PGCs. These differentially methylated regions may be directly or indirectly associated with gene expression during early embryonic development, and the epigenetic difference could be evolutionally conserved between mammals and birds.

Female gender and pulmonary arterial hypertension: a complex relationship

Pulmonary arterial hypertension (PAH) is a severe clinical condition defined as mean pulmonary artery pressure ≥25 mmHg and normal pulmonary capillary wedge pressure (≤15 mmHg). In PAH the increase in pulmonary pressure is due to an intrinsic disease of the small pulmonary arteries (resistance vessels) characterized by vascular proliferation and remodeling. The increase in pulmonary vascular resistance with subsequent elevation of the right ventricular afterload leads to right ventricular failure after variable periods of time. Although targeted disease therapies have been developed over the last decade that resulted in improved quality of life and outcome for PAH patients, the prognosis is still severe and there remains no cure for this disease. From a clinical standpoint, PAH includes a group of heterogeneous pathological conditions: in idiopathic, heritable and drug- and toxin-induced PAH, since there are no predisposing clinical conditions, the structural changes in pulmonary circulation are "isolated"; on the other hand, PAH may be associated with some predisposing diseases such as connective tissue disease, HIV infection, portal hypertension, congenital heart disease, schistosomiasis, and chronic hemolytic anemia. PAH can affect individuals of all age groups, and mean age at diagnosis is around 50 years. Epidemiological data show a great preponderance of females in PAH; the high prevalence of females is particularly evident in the so-called "isolated" PAH forms, whereas in PAH associated with other diseases the female:male ratio is strongly influenced by the epidemiological features of the specific predisposing condition. The reason for the higher female prevalence in PAH has never been clarified: some hypotheses involve the role of sexual hormones (estrogens), autoimmunity, or an X-linked locus in disease predisposition. Female gender is not associated with a different clinical presentation. However, the age of onset tends to be earlier in females than in males. As far as prognosis is concerned, male patients are at greater risk of mortality despite similar therapeutic management: this could be linked to the influence of sexual hormones, which may favor disease development but may also have favorable effects on outcome, especially on treatment response (the "estrogen paradox"). The possibility that female sex hormones may influence the development and clinical expression of PAH is underlined by the frequent onset of PAH during pregnancy or shortly after delivery, and by the description of obligate carriers of heritable PAH (BMPR2 mutation carriers) in whom hormone replacement therapy seems to trigger the manifestation of the disease. Nevertheless, a definite pathobiological mechanism to explain the association between female sex hormones and PAH has yet to be identified. The link between pregnancy and PAH is also uncertain. However, the hemodynamic changes that occur during gestation, and mainly during labor and delivery, are poorly tolerated by PAH patients: this is reflected in the timing of clinical deterioration and the increased mortality in the late stages of pregnancy when maximum increase in blood volume and cardiac output occurs. As such, the current clinical recommendation is that pregnancy should strongly be discouraged and if it occurs, early termination is advised. When PAH is not diagnosed until late in pregnancy, close follow-up of the mother is mandatory and elective planned delivery is recommended: care of pregnant women with PAH requires a highly planned, multidisciplinary approach, involving obstetricians, pulmonary hypertension specialists, anesthesiologists and intensivists, preferably in a dedicated PAH referral center. Use of female hormones for birth control and postmenopausal replacement therapy in PAH patients still remains controversial; in fact, although it has been suspected to be a trigger factor for PAH development, a formal association based on case-control studies has not been documented.

Model‐based comparisons of phylogeographic scenarios resolve the intraspecific divergence of cactophilic Drosophila mojavensis

The cactophilic fly Drosophila mojavensis exhibits considerable intraspecific genetic structure across allopatric geographic regions and shows associations with different host cactus species across its range. The divergence between these populations has been studied for more than 60years, yet their exact historical relationships have not been resolved. We analysed sequence data from 15 intronic X-linked loci across populations from Baja California, mainland Sonora-Arizona and Mojave Desert regions under an isolation-with-migration model to assess multiple scenarios of divergence. We also compared the results with a pre-existing sequence data set of eight autosomal loci. We derived a population tree with Baja California placed at its base and link their isolation to Pleistocene climatic oscillations. Our estimates suggest the Baja California population diverged from an ancestral Mojave Desert/mainland Sonora-Arizona group around 230,000-270,000years ago, while the split between the Mojave Desert and mainland Sonora-Arizona populations occurred one glacial cycle later, 117,000-135,000years ago. Although we found these three populations to be effectively allopatric, model ranking could not rule out the possibility of a low level of gene flow between two of them. Finally, the Mojave Desert population showed a small effective population size, consistent with a historical population bottleneck. We show that model-based inference from multiple loci can provide accurate information on the historical relationships of closely related groups allowing us to set into historical context a classic system of incipient ecological speciation.

Hand preference in humans is associated with testosterone levels and androgen receptor gene polymorphism

Exposure of the central nervous system to androgens during the early developmental period has been proposed to play a role in the establishment of hand preference in males. Existing data, however, are inconclusive. In the present investigation, handedness was assessed in a large sample of left-, mixed-, and right-handed men (N=180) using a standardized handedness inventory. Saliva sampling was used to assay levels of bioavailable testosterone and DNA genotyping was carried out to quantify AR-CAG repeat length, a genetic marker of the capacity of the androgen receptor to respond to testosterone. Strongly left-handed males were found to have lower levels of bioavailable testosterone than right-handed males, while males with mixed handedness exhibited a weaker androgen receptor, but no significant difference from right-handers in circulating testosterone levels. These findings support the view that testosterone could play a role in the development of hand preference in males. Furthermore, because the AR gene lies on the X chromosome, it provides a potential theoretical bridge to genetic theories of handedness that postulate the existence of an X-linked locus important in the establishment of hand preference.

The Role of Background Selection in Shaping Patterns of Molecular Evolution and Variation: Evidence from Variability on the Drosophila X Chromosome

In the putatively ancestral population of Drosophila melanogaster, the ratio of silent DNA sequence diversity for X-linked loci to that for autosomal loci is approximately one, instead of the expected "null" value of 3/4. One possible explanation is that background selection (the hitchhiking effect of deleterious mutations) is more effective on the autosomes than on the X chromosome, because of the lack of crossing over in male Drosophila. The expected effects of background selection on neutral variability at sites in the middle of an X chromosome or an autosomal arm were calculated for different models of chromosome organization and methods of approximation, using current estimates of the deleterious mutation rate and distributions of the fitness effects of deleterious mutations. The robustness of the results to different distributions of fitness effects, dominance coefficients, mutation rates, mapping functions, and chromosome size was investigated. The predicted ratio of X-linked to autosomal variability is relatively insensitive to these variables, except for the mutation rate and map length. Provided that the deleterious mutation rate per genome is sufficiently large, it seems likely that background selection can account for the observed X to autosome ratio of variability in the ancestral population of D. melanogaster. The fact that this ratio is much less than one in D. pseudoobscura is also consistent with the model's predictions, since this species has a high rate of crossing over. The results suggest that background selection may play a major role in shaping patterns of molecular evolution and variation.

Impact of Genetic Architecture on the Relative Rates of X versus Autosomal Adaptive Substitution

Molecular evolutionary theory predicts that the ratio of autosomal to X-linked adaptive substitution (K(A)/K(x)) is primarily determined by the average dominance coefficient of beneficial mutations. Although this theory has profoundly influenced analysis and interpretation of comparative genomic data, its predictions are based upon two unverified assumptions about the genetic basis of adaptation. The theory assumes that 1) the rate of adaptively driven molecular evolution is limited by the availability of beneficial mutations, and 2) the scaling of evolutionary parameters between the X and the autosomes (e.g., the beneficial mutation rate, and the fitness effect distribution of beneficial alleles, per X-linked versus autosomal locus) is constant across molecular evolutionary timescales. Here, we show that the genetic architecture underlying bouts of adaptive substitution can influence both assumptions, and consequently, the theoretical relationship between K(A)/K(x) and mean dominance. Quantitative predictions of prior theory apply when 1) many genomically dispersed genes potentially contribute beneficial substitutions during individual steps of adaptive walks, and 2) the population beneficial mutation rate, summed across the set of potentially contributing genes, is sufficiently small to ensure that adaptive substitutions are drawn from new mutations rather than standing genetic variation. Current research into the genetic basis of adaptation suggests that both assumptions are plausibly violated. We find that the qualitative positive relationship between mean dominance and K(A)/K(x) is relatively robust to the specific conditions underlying adaptive substitution, yet the quantitative relationship between dominance and K(A)/K(x) is quite flexible and context dependent. This flexibility may partially account for the puzzlingly variable X versus autosome substitution patterns reported in the empirical evolutionary genomics literature. The new theory unites the previously separate analysis of adaptation using new mutations versus standing genetic variation and makes several useful predictions about the interaction between genetic architecture, evolutionary genetic constraints, and effective population size in determining the ratio of adaptive substitution between autosomal and X-linked genes.

Genetic Association of Multiple Sclerosis with the Marker rs391745 near the Endogenous Retroviral Locus HERV-Fc1: Analysis of Disease Subtypes

We have previously described the occurrence of multiple sclerosis (MS) to be associated with human endogenous retroviruses, specifically the X-linked viral locus HERV-Fc1. The aim of this study was to investigate a possible association of the HERV-Fc1 locus with subtypes of MS. MS patients are generally subdivided into three categories: Remitting/Relapsing and Secondary Progressive, which together constitute Bout Onset MS, and Primary Progressive. In this study of 1181 MS patients and 1886 controls we found that Bout Onset MS was associated with the C-allele of the marker rs391745 near the HERV-Fc1 locus (p = 0.003), while primary progressive disease was not. The ability to see genetic differences between subtypes of MS near this gene speaks for the involvement of the virus HERV-Fc1 locus in modifying the disease course of MS.

G6PD deficiency and malaria selection

In his valuable review on genetics of malaria resistance, Hedrick (2011) tackles the interesting issue of the protection against Plasmodium falciparum afforded by G6PD deficiency. Specifically, he quotes data from a study (Ruwende et al., 1995) conducted in Gambia and in Kenya, from which it was claimed that the degree of protection was similar in hemizygous G6PD-deficient males and in heterozygous G6PD-deficienct females. Hedrick (2011) points out that this is rather paradoxical, because it would lead to fixation of the G6PD-deficient allele(s), which does not seem to have taken place; and he reproduces from that paper a figure that offers an explanation on the basis of an ad-hoc hypothesis. Hedrick (2011) has not made mention of a previous study of 699 children (Bienzle et al., 1972) in which, based on the distribution of P. falciparum parasitaemia values in children in Southwestern Nigeria, we reported that there was significant protection of girls heterozygous for G6PD deficiency, whereas we did not find the same in hemizygous G6PD-deficient boys. Thus, these data are fully compatible with balanced polymorphism at or near equilibrium. Although this is not mentioned in Hedrick's review, it has now been found (Clark et al., 2009) that the most common G6PD deficiency allele in the Gambia is G6PD A− having the two mutations N126D and L323P, and not G6PD A− having the two mutations N126D and M68V. Although both of these alleles were known since 1989 (Beutler et al., 1989), only the latter was tested for in the study by Ruwende et al. (1995) on the assumption that the frequency of the former would be negligible. As a result, we can estimate that in the Gambia component of the study, about two-thirds of the G6PD-deficient samples were misclassified as normal; this leaves the Kenya component of the study, consisting of a group of 935 children, within which the differences observed did not reach statistical significance. It is not clear why the paper by Bienzle et al. (1972) was ignored. If the reason was because it did not include molecular analysis (the G6PD gene was not cloned until several years later: Persico et al., 1986), this was not a good reason, because robust genotypic classification was obtained in that study by combining no less than three independent methodologies. The reliability of our genotypic classification was confirmed by the fact that the data were in Hardy–Weinberg equilibrium. It is paradoxical that the review missed this study, which had incorporated deliberate safeguards against genotype misclassification; and if more than one type of G6PD A− had been present in the population sample, this would not have affected the results, because by the methods used, all types of G6PD A− were identified. Bennett (1958) pointed out that increased fitness of heterozygotes is not in itself either necessary or sufficient for reaching equilibrium under selection at an X-linked locus; this depends on the fitness values of the other genotypes. However, in the case of G6PD deficiency, there is every reason to assume that the fitness of normal male hemizygotes is the same as that of normal female homozygotes, and that the fitness of G6PD-deficient male hemizygotes is the same as that of G6PD-deficient female homozygotes. Under these conditions, it has been pointed out by Haldane and Jayakar (1964) and by Kirkman (1966) that the female heterozygote must be the genotype with the highest fitness. There is no a priori certainty that, in any particular population, the G6PD polymorphism must be at equilibrium. In principle, malaria selection could ultimately cause fixation of a G6PD deficiency allele. However, this is not very likely, because (a) this has not happened in any population, even with holoendemic malaria as in Nigeria, and (b) both P. falciparum malaria endemicity and G6PD deficiency A− are estimated (Tishkoff et al., 2001) to be at least 5000 years old (corresponding to some 200 generations). To obtain evidence on genetic protection against malaria is not easy, in part because the differences in fitness associated with individual genotypes may be small; therefore, it is probably best to consider all independent studies (for instance, in addition to those already mentioned, that by Guindo et al. (2007). At the present state of knowledge, two independent studies (Bienzle et al., 1972; Clark et al., 2009) in two different populations, nearly 40 years apart, are consistent, with G6PD deficiency A− being a balanced polymorphism with heterozygote advantage.

Methylation of L1Hs promoters is lower on the inactive X, has a tendency of being higher on autosomes in smaller genomes and shows inter-individual variability at some loci

LINE-1 repeats account for ∼17% of the human genome. Little is known about their individual methylation patterns, because their repetitive, almost identical sequences make them difficult to be individually targeted. Here, we used bisulfite conversion to study methylation at individual LINE-1 repeats. The loci studied included 39 X-linked loci and 5 autosomal loci. On the X chromosome in women, we found statistically significant less methylation at almost all L1Hs compared with men. Methylation at L1P and L1M did not correlate with the inactivation status of the host DNA, while the majority of L1Hs that were possible to be studied lie in inactivated regions. To investigate whether the male–female differences at L1Hs on the X are linked to the inactivation process itself rather than to a mere influence of gender, we analyzed six of the L1Hs loci on the X chromosome in Turners and Klinefelters which have female and male phenotype, respectively, but with reversed number of X chromosomes. We could confirm that all samples with two X chromosomes are hypomethylated at the L1Hs loci. Therefore, the inactive X is hypomethylated at L1Hs; the latter could play an exclusive role in the X chromosome inactivation process. At autosomal L1Hs, methylation levels showed a correlation tendency between methylation level and genome size, with higher methylation observed at most loci in individuals with one X chromosome and the lowest in XXY individuals. In summary, loci-specific LINE-1 methylation levels show considerable plasticity and depend on genomic position and constitution.

Evidence for X‐linked introgression between molecular forms of Anopheles gambiae from Angola

The M and S molecular forms of the African malaria vector Anopheles gambiae (Diptera: Culicidae) are morphologically identical incipient species in which reproductive isolation is incomplete, enabling low-level gene flow between forms. In an attempt to find differences between the M and S forms, sequence variation was studied at loci along the X chromosome in adult female An. gambiae from Angola. A high proportion of M form specimens from Angola (79% of the 456 X chromosomes sampled) were found to contain a 16-bp insertion in intron 4 of the X-linked GPRCCK1 locus, relative to the AgamP3 release of the An. gambiae PEST genome sequence. The insertion was in Hardy-Weinberg equilibrium in Angolan M form populations. The same insertion was found in all S form specimens examined, regardless of where in Africa they were sampled, but was absent from a sample of M form specimens collected in Ghana, Bioko and Mali. In M form specimens from Angola, there was an association between alleles at the GPRCCK1 locus and those at a microsatellite locus, AGXH678, close to the centromere of the X chromosome, with significant linkage disequilibrium between loci separated by 0.472 Mbp (P < 0.033). We show that the insertion results from introgression from the S form into the M form, rather than from the retention of an ancestral character. Gene flow from the S to M form could allow genes of adaptive value to be transferred, including those conferring insecticide resistance and others influencing ecology and behaviour, and thus malaria transmission and control. We discuss factors that may have led to this introgression event.

Refinement of the X-linked Nonsyndromic High-Grade Myopia LocusMYP1on Xq28 and Exclusion of 13 Known Positional Candidate Genes by Direct Sequencing

Myopia is a common vision problem affecting almost one third of the world's population. It can occur as an isolated genetic condition or be associated with other anomalies and/or syndromes. Seventeen myopia loci have been identified on various chromosomes; however, no specific gene mutations have yet been identified. Two large multigeneration Asian Indian pedigrees (UR006 and UR077) with isolated, nonsyndromic myopia were studied, in which the condition appeared to segregate as an X-linked recessive trait (MYP1; MIM 310460). The degree of myopia was variable in both families, ranging from -6 to -23 D (mean, -8.48 D) with the majority >7.0 D. To map the myopia locus in these families, polymorphic microsatellite markers covering the entire X chromosome were used in linkage analyses performed on 42 genomic DNA samples (13 affected and 29 normal) from both families. Marker DXYS154, which is located within the pseudoautosomal region in distal Xq28 (PAR2; pseudoautosomal region 2), gave a combined maximum LOD score of 5.3 at = 0 under an autosomal recessive model. Other markers in the region (near but not within the PAR2 region) that showed no recombination with the phenotype in both the families included DXS1108, DXS8087, and F8i13. Observation of recombination in family UR006 refined the disease locus to a ∼1.25-Mb region flanked by the proximal marker DXS1073 and distal marker DXYS154. Mutation search in exons and splice junctions of candidate genes CTAG2, GAB3, MPP1, F8Bver, FUNDC2, VBP1, RAB39B, CLIC2, TMLHE, SYBL, IL9R, SPRY3, and CXYorf1 did not detect a pathogenic or predisposing variant.

Genetic dissection of testis weight in mice: quantitative trait locus analysis using F2 intercrosses between strains with extreme testis weight, and association study using Y-consomic strains

In the present study, dissection of genetic bases of testis weight in mice was performed. Autosomes and the X chromosome were searched using traditional quantitative trait locus (QTL) scans, and the Y chromosome was searched by association studies of Y-consomic strains. QTL analysis was performed in ♀DDD×♂CBA F(2) mice; the inbred mouse DDD has the heaviest testes, whereas the inbred mouse CBA has the lightest testes. Two significant testis weight QTLs were identified on chromosomes 1 and X. A DDD allele was associated with increased and decreased testis weight at the locus on chromosomes 1 and X, respectively. In the reciprocal cross ♀CBA×♂DDD F(2) mice, QTL on chromosome 1, and not on chromosome X, had a significant effect on testis weight. The DDD allele at the X-linked locus could not sustain testis weight in combination with the Y chromosome of the CBA strain. The Y chromosome per se had a significant effect on testis weight, i.e., DH-Chr Y(DDD) had significantly heavier testes than DH-Chr Y(CBA). On the basis of the results of Y-chromosome-wide association studies using 17 Y-consomic strains, variations in Uty, Usp9y, and Sry were significantly associated with testis weight. Thus, testis weight is a complex quantitative phenotype controlled by multiple genes on autosomes and sex chromosomes and their interactions.