Modern horses represent heterogeneous populations specifically selected for appearance and performance. Genomic regions under high selective pressure show characteristic runs of homozygosity (ROH) ...which represent a low genetic diversity. This study aims at detecting the number and functional distribution of ROHs in different horse populations using next generation sequencing data.
Next generation sequencing was performed for two Sorraia, one Dülmen Horse, one Arabian, one Saxon-Thuringian Heavy Warmblood, one Thoroughbred and four Hanoverian. After quality control reads were mapped to the reference genome EquCab2.70. ROH detection was performed using PLINK, version 1.07 for a trimmed dataset with 11,325,777 SNPs and a mean read depth of 12. Stretches with homozygous genotypes of >40 kb as well as >400 kb were defined as ROHs. SNPs within consensus ROHs were tested for neutrality. Functional classification was done for genes annotated within ROHs using PANTHER gene list analysis and functional variants were tested for their distribution among breed or non-breed groups.
ROH detection was performed using whole genome sequences of ten horses of six populations representing various breed types and non-breed horses. In total, an average number of 3492 ROHs were detected in windows of a minimum of 50 consecutive homozygous SNPs and an average number of 292 ROHs in windows of 500 consecutive homozygous SNPs. Functional analyses of private ROHs in each horse revealed a high frequency of genes affecting cellular, metabolic, developmental, immune system and reproduction processes. In non-breed horses, 198 ROHs in 50-SNP windows and seven ROHs in 500-SNP windows showed an enrichment of genes involved in reproduction, embryonic development, energy metabolism, muscle and cardiac development whereas all seven breed horses revealed only three common ROHs in 50-SNP windows harboring the fertility-related gene YES1. In the Hanoverian, a total of 18 private ROHs could be shown to be located in the region of genes potentially involved in neurologic control, signaling, glycogen balance and reproduction. Comparative analysis of homozygous stretches common in all ten horses displayed three ROHs which were all located in the region of KITLG, the ligand of KIT known to be involved in melanogenesis, haematopoiesis and gametogenesis.
The results of this study give a comprehensive insight into the frequency and number of ROHs in various horses and their potential influence on population diversity and selection pressures. Comparisons of breed and non-breed horses suggest a significant artificial as well as natural selection pressure on reproduction performance in all types of horse populations.
Chronic obstructive pulmonary disease (COPD) is induced by cigarette smoking and characterized by inflammation of airway tissue. Since smokers with COPD have a higher risk of developing lung cancer ...than those without, we hypothesized that they carry more mutations in affected tissue. We called somatic mutations in airway brush samples from medium-coverage whole genome sequencing data from healthy never and ex-smokers (n = 8), as well as from ex-smokers with variable degrees of COPD (n = 4). Owing to the limited concordance of resulting calls between the applied tools we built a consensus, a strategy that was validated with high accuracy for cancer data. However, consensus calls showed little promise of representing true positives due to low mappability of corresponding sequence reads and high overlap with positions harbouring known genetic polymorphisms. A targeted re-sequencing approach suggested that only few mutations would survive stringent verification testing and that our data did not allow the inference of any difference in the mutational load of bronchial brush samples between former smoking COPD cases and controls. High polyclonality in airway brush samples renders medium-depth sequencing insufficient to provide the resolution to detect somatic mutations. Deep sequencing data of airway biopsies are needed to tackle the question.
Stimulation of bone formation is a key therapeutic target in osteoporosis. Runx2 is a runt domain transcription factor essential to osteoblast differentiation, bone remodeling, and fracture healing. ...Runx2 knockout mice exhibit a complete lack of ossification, while overexpression of this gene in transgenic mice results in an osteoporotic phenotype. Thus, RUNX2 is a good candidate for the genetic determination of osteoporosis. In this association study, the effects of the -330 G/T polymorphism in promoter 1 and the -1025 T/C polymorphism (rs7771980) in promoter 2 of RUNX2 were tested in relation to lumbar spine (LS) and femoral neck (FN) bone mineral density (BMD) in a cohort of 821 Spanish postmenopausal women. The minor allele frequencies for the two polymorphisms were 0.15 and 0.07, respectively. The two polymorphisms, located more than 90 kb apart, were not in linkage disequilibrium (D' = 0.27, r (2) = 0.028). In an ANCOVA test adjusting by weight, height, age, and years since menopause, the -330 G/T polymorphism was not associated with any of the phenotypes analyzed, while we found the -1025 T/C polymorphism to be associated with FN BMD (p = 0.001). In particular, individuals carrying the TC genotype had higher mean adjusted FN BMD values than those bearing the TT genotype. Our results highlight the importance of this RUNX2 promoter 2 polymorphism in FN BMD determination.
Mutations in the low-density lipoprotein receptor-related protein 5 (LRP5) gene cause rare syndromes characterized by altered bone mineral density (BMD). More common LRP5 variants may affect ...osteoporosis risk in the general population.
To generate large-scale evidence on whether 2 common variants of LRP5 (Val667Met, Ala1330Val) and 1 variant of LRP6 (Ile1062Val) are associated with BMD and fracture risk.
Prospective, multicenter, collaborative study of individual-level data on 37,534 individuals from 18 participating teams in Europe and North America. Data were collected between September 2004 and January 2007; analysis of the collected data was performed between February and May 2007. Bone mineral density was assessed by dual-energy x-ray absorptiometry. Fractures were identified via questionnaire, medical records, or radiographic documentation; incident fracture data were available for some cohorts, ascertained via routine surveillance methods, including radiographic examination for vertebral fractures.
Bone mineral density of the lumbar spine and femoral neck; prevalence of all fractures and vertebral fractures.
The Met667 allele of LRP5 was associated with reduced lumbar spine BMD (n = 25,052 number of participants with available data; 20-mg/cm2 lower BMD per Met667 allele copy; P = 3.3 x 10(-8)), as was the Val1330 allele (n = 24,812; 14-mg/cm2 lower BMD per Val1330 copy; P = 2.6 x 10(-9)). Similar effects were observed for femoral neck BMD, with a decrease of 11 mg/cm2 (P = 3.8 x 10(-5)) and 8 mg/cm2 (P = 5.0 x 10(-6)) for the Met667 and Val1330 alleles, respectively (n = 25 193). Findings were consistent across studies for both LRP5 alleles. Both alleles were associated with vertebral fractures (odds ratio OR, 1.26; 95% confidence interval CI, 1.08-1.47 for Met667 2001 fractures among 20 488 individuals and OR, 1.12; 95% CI, 1.01-1.24 for Val1330 1988 fractures among 20,096 individuals). Risk of all fractures was also increased with Met667 (OR, 1.14; 95% CI, 1.05-1.24 per allele 7876 fractures among 31,435 individuals)) and Val1330 (OR, 1.06; 95% CI, 1.01-1.12 per allele 7802 fractures among 31 199 individuals). Effects were similar when adjustments were made for age, weight, height, menopausal status, and use of hormone therapy. Fracture risks were partly attenuated by adjustment for BMD. Haplotype analysis indicated that Met667 and Val1330 variants both independently affected BMD. The LRP6 Ile1062Val polymorphism was not associated with any osteoporosis phenotype. All aforementioned associations except that between Val1330 and all fractures and vertebral fractures remained significant after multiple-comparison adjustments.
Common LRP5 variants are consistently associated with BMD and fracture risk across different white populations. The magnitude of the effect is modest. LRP5 may be the first gene to reach a genome-wide significance level (a conservative level of significance herein, unadjusted P < 10(-7) that accounts for the many possible comparisons in the human genome) for a phenotype related to osteoporosis.
LRP5 encodes the low‐density lipoprotein receptor‐related protein 5, a transmembrane protein involved in Wnt signaling. LRP5 is an important regulator of osteoblast growth and differentiation, ...affecting bone mass in vertebrates. Whether common variations in LRP5 are associated with normal BMD variation or osteoporotic phenotypes is of great relevance. We used a haplotype‐based approach to search for common disease‐associated variants in LRP5 in a cohort of 964 Spanish postmenopausal women. Twenty‐four SNPs were selected, covering the LRP5 region, including the missense changes p.V667M and p.A1330V. The SNPs were genotyped and evaluated for association with BMD at the lumbar spine (LS) or femoral neck (FN) and with osteoporotic fracture, at single SNP and haplotype levels, by regression methods. Association with LS BMD was found for SNP 1, rs312009, located in the 5′‐flanking region (p = 0.011, recessive model). SNP 6, rs2508836, in intron 1, was also associated with BMD, both at LS (p = 0.025, additive model) and FN (p = 0.031, recessive model). Two polymorphisms were associated with fracture: SNP 11, rs729635, in intron 1, and SNP 15, rs643892, in intron 5 (p = 0.007 additive model and p = 0.019 recessive model, respectively). Haplotype analyses did not provide additional information, except for haplotype “GC” of the block located at the 3′end of the gene. This haplotype spans intron 22 and the 3′ untranslated region and was associated with FN BMD (p = 0.029, one copy of the haplotype versus none). In silico analyses showed that SNP 1 (rs312009) lies in a putative RUNX2 binding site. Electro‐mobility shift assays confirmed RUNX2 binding to this site.
Osteoporosis is a complex disease involving many putative genetic factors. Association analysis of functional SNPs in candidate genes is an important tool for their identification. However, this ...approach is affected by limited power, population stratification, and other drawbacks that lead to discordant results. Replication in independent cohorts is essential. We performed association analyses of three functional polymorphisms previously associated with bone phenotypes—namely, Ala222Val in
MTHFR
, Ile1062Val in
LRP6
, and −13910C>T in
LCT
—in a cohort of 944 postmenopausal Spanish women, all of them with lumbar spine (LS) bone mineral density (BMD) data and most with femoral neck (FN) BMD and fracture data. We found significant differences between genotypes only for the
MTHFR
polymorphism and vertebral factures, with an OR of 2.27 (95% CI 1.17–4.38) for the TT vs. CC/CT genotypes,
P
= 0.018. We present genotype and allele frequency data for
LCT
−13910C>T for a Spanish population, where the T allele (conferring lactase persistence) has a frequency of 38.6%. Genotype frequencies were consistent with observed clines in Europe and with the prevalence of lactase nonpersistence. The
LCT
−13910C>T polymorphism was significantly associated with height and weight, such that T allele carriers were 0.88 cm taller (95% CI 0.08–1.59 cm,
P
= 0.032, adjusted by age) than CC individuals and TT homozygotes were 1.91 kg heavier than CC/CT individuals (95% CI 0.11–3.71 kg,
P
= 0.038, adjusted by age). In conclusion, no significant association was observed between the studied polymorphisms and LS BMD or FN BMD in postmenopausal Spanish women, and only
MTHFR
Ala222Val was associated with vertebral fractures.
Most great ape genetic variation remains uncharacterized; however, its study is critical for understanding population history, recombination, selection and susceptibility to disease. Here we sequence ...to high coverage a total of 79 wild- and captive-born individuals representing all six great ape species and seven subspecies and report 88.8 million single nucleotide polymorphisms. Our analysis provides support for genetically distinct populations within each species, signals of gene flow, and the split of common chimpanzees into two distinct groups: Nigeria-Cameroon/western and central/eastern populations. We find extensive inbreeding in almost all wild populations, with eastern gorillas being the most extreme. Inferred effective population sizes have varied radically over time in different lineages and this appears to have a profound effect on the genetic diversity at, or close to, genes in almost all species. We discover and assign 1,982 loss-of-function variants throughout the human and great ape lineages, determining that the rate of gene loss has not been different in the human branch compared to other internal branches in the great ape phylogeny. This comprehensive catalogue of great ape genome diversity provides a framework for understanding evolution and a resource for more effective management of wild and captive great ape populations.
Analyzing the DNA methylome of multiple myeloma (MM), a plasma cell neoplasm, by whole-genome bisulfite sequencing and high-density arrays, we observed regional DNA hypermethylation embedded in ...extensive global hypomethylation. In contrast to the widely reported DNA hypermethylation of promoter-associated CpG islands (CGIs) in cancer, hypermethylated sites in MM as compared to normal plasma cells were located outside CpG islands and were unexpectedly associated with intronic enhancer regions active in normal B cells. Both RNA-seq and in vitro reporter assays indicated that enhancer hypermethylation is globally associated with downregulation of its host genes. ChIP-seq and DNAseI-seq further revealed that DNA hypermethylation in these regions was related to enhancer decommissioning. Hypermethylated enhancer regions overlap with binding sites of B-cell specific transcription factors (TFs) and the degree of enhancer methylation inversely correlated with expression levels of these TFs in MM. Furthermore, hypermethylated regions in MM were methylated in stem cells and gradually became demethylated during normal B-cell differentiation suggesting that MM cells reacquire epigenetic features of undifferentiated cells upon loss of expression of B-cell specific TFs. Overall, we have identified DNA hypermethylation of developmentally-regulated enhancers as a new type of epigenetic modification associated with the pathogenesis of MM.
No relevant conflicts of interest to declare.
Introduction: Modulation of the DNA methylation landscape during cell differentiation is a well-established phenomenon. The B-cell lineage represents a paradigmatic cellular model to study the ...dynamic epigenome during cell development and specification because major B-cell maturation stages are well defined and display differential phenotypic and gene expression features. Furthermore, different B-cell subpopulations show different proliferation abilities, microenvironmental influences and life spans, providing a window of opportunity to study the epigenome in the context of multiple processes.
Methods: We performed whole-genome bisulfite sequencing (WGBS), high-density methylation microarrays and gene expression profiling of ten purified human B-cell subpopulations spanning the entire differentiation program, ranging from uncommitted progenitors to terminally-differentiated plasma cells.
Results: The results of both WGBS and methylation microarrays indicate that B-cell ontogenesis involves an extensive and gradual reconfiguration of the DNA methylome. We uncovered that non-CpG methylation at CpApC trinucleotides is present in progenitor cells and disappears upon B-cell commitment independently of CpG demethylation. CpG methylation, in contrast, changed extensively during the entire B-cell maturation program, with one quarter of all measured CpGs showing dynamic methylation. B-cell enhancers suffered more extensive methylation changes than promoter regions, especially in the early differentiation steps up to the germinal center B-cell (gcBC) stage, and their demethylation seemed to be mediated by binding of lineage-specific transcription factors. Enhancers with dynamic methylation were related to genes involved in a large B-cell network that showed high gene expression variability throughout differentiation. In highly proliferative gcBCs, we observed a shift of dynamic methylation from regulatory towards non-functional elements; gcBCs start to undergo global demethylation of late-replicating heterochromatic regions and methylation of polycomb-repressed regions. This signature becomes particularly extensive in long-lived memory B cells and plasma cells, indicating that these changes start in highly proliferative cells and then accumulate in non-proliferative cells with extended lifespan.
Conclusion: Our epigenomic analysis of the B-cell differentiation program extends our knowledge on how the DNA methylome is modulated during cell specification and maturation and offers a resource for researchers in the field, both at global and single gene levels.
No relevant conflicts of interest to declare.
PDF
