Buccals are likely to be a more informative surrogate tissue than blood for epigenome-wide association studies

There is increasing evidence that interindividual epigenetic variation is an etiological factor in common human diseases. Such epigenetic variation could be genetic or non-genetic in origin, and epigenome-wide association studies (EWASs) are underway for a wide variety of diseases/phenotypes. However, performing an EWAS is associated with a range of issues not typically encountered in genome-wide association studies (GWASs), such as the tissue to be analyzed. In many EWASs, it is not possible to analyze the target tissue in large numbers of live humans, and consequently surrogate tissues are employed, most commonly blood. But there is as yet no evidence demonstrating that blood is more informative than buccal cells, the other easily accessible tissue. To assess the potential of buccal cells for use in EWASs, we performed a comprehensive analysis of a buccal cell methylome using whole-genome bisulfite sequencing. Strikingly, a buccal vs. blood comparison reveals > 6X as many hypomethylated regions in buccal. These tissue-specific differentially methylated regions (tDMRs) are strongly enriched for DNaseI hotspots. Almost 75% of these tDMRs are not captured by commonly used DNA methylome profiling platforms such as Reduced Representational Bisulfite Sequencing and the Illumina Infinium HumanMethylation450 BeadChip, and they also display distinct genomic properties. Buccal hypo-tDMRs show a statistically significant enrichment near SNPs associated to disease identified through GWASs. Finally, we find that, compared with blood, buccal hypo-tDMRs show significantly greater overlap with hypomethylated regions in other tissues. We propose that for non-blood based diseases/phenotypes, buccal will be a more informative tissue for EWASs.     

Identification of type 1 diabetes-associated DNA methylation variable positions that precede disease diagnosis

Monozygotic (MZ) twin pair discordance for childhood-onset Type 1 Diabetes (T1D) is ∼50%, implicating roles for genetic and non-genetic factors in the aetiology of this complex autoimmune disease. Although significant progress has been made in elucidating the genetics of T1D in recent years, the non-genetic component has remained poorly defined. We hypothesized that epigenetic variation could underlie some of the non-genetic component of T1D aetiology and, thus, performed an epigenome-wide association study (EWAS) for this disease. We generated genome-wide DNA methylation profiles of purified CD14⁺ monocytes (an immune effector cell type relevant to T1D pathogenesis) from 15 T1D–discordant MZ twin pairs. This identified 132 different CpG sites at which the direction of the intra-MZ pair DNA methylation difference significantly correlated with the diabetic state, i.e. T1D–associated methylation variable positions (T1D–MVPs). We confirmed these T1D–MVPs display statistically significant intra-MZ pair DNA methylation differences in the expected direction in an independent set of T1D–discordant MZ pairs (P = 0.035). Then, to establish the temporal origins of the T1D–MVPs, we generated two further genome-wide datasets and established that, when compared with controls, T1D–MVPs are enriched in singletons both before (P = 0.001) and at (P = 0.015) disease diagnosis, and also in singletons positive for diabetes-associated autoantibodies but disease-free even after 12 years follow-up (P = 0.0023). Combined, these results suggest that T1D–MVPs arise very early in the etiological process that leads to overt T1D. Our EWAS of T1D represents an important contribution toward understanding the etiological role of epigenetic variation in type 1 diabetes, and it is also the first systematic analysis of the temporal origins of disease-associated epigenetic variation for any human complex disease.     

An assessment of plant growth and N<Subscript>2</Subscript> fixation in soybean genotypes grown in uninoculated soils collected from different locations in Ethiopia

Pinus heldreichii H. Christ. is a tertiary relict and endemic to the western Balkans and southern part of Apennine peninsula. It is an Oro-Mediterranean species occurring at altitudes between 1200 and 2000 m and primarily on calcareous soils. P. heldreichii forests are of key importance for nature conservation, protection against gravitational natural hazards, landscape conservation and recreation. However, these forests are currently highly fragmented and require the elaboration of guidelines for sustainable management and conservation that should be based on scientific knowledge. Ectomycorrhizal (ECM) fungi are important for successful regeneration, establishment and growth of P. heldreichii. The aim of this study was to investigate ECM and other fungal communities associated with fine roots of P. heldreichii at two different sites in Ku?i Mountains, south-eastern Montenegro. Roots and soil were sampled from 70 trees. Soil was subjected to chemical analyses, fine roots were morphotyped and selected root morphotypes were Sanger sequenced using ITS rDNA as a marker. Sequencing resulted in 431 high-quality sequences representing 147 different fungal species including a large number of ECM species. The most common species were ECM fungi Lactarius sanguifluus (5.1%), Wilcoxina rehmii (4.2%) and Amphinema sp. KK28 (3.2%). Climatic factors were similar between the sites, but site size, inclination, elevation, tree age (old growth versus young trees), and some soil characteristics differed. The results demonstrate relatively high fungal diversity and site-specific effects on abundance and composition of fungal communities in fine roots of P. heldreichii growing in high-altitude marginal habitats.