Regulation of DNA Methylation Patterns by CK2-Mediated Phosphorylation of Dnmt3a

DNA methylation is a central epigenetic modification that is established by de novo DNA methyltransferases. The mechanisms underlying the generation of genomic methylation patterns are still poorly understood. Using mass spectrometry and a phosphospecific Dnmt3a antibody, we demonstrate that CK2 phosphorylates endogenous Dnmt3a at two key residues located near its PWWP domain, thereby downregulating the ability of Dnmt3a to methylate DNA. Genome-wide DNA methylation analysis shows that CK2 primarily modulates CpG methylation of several repeats, most notably of Alu SINEs. This modulation can be directly attributed to CK2-mediated phosphorylation of Dnmt3a. We also find that CK2-mediated phosphorylation is required for localization of Dnmt3a to heterochromatin. By revealing phosphorylation as a mode of regulation of de novo DNA methyltransferase function and by uncovering a mechanism for the regulation of methylation at repetitive elements, our results shed light on the origin of DNA methylation patterns

Statistical Inference of In Vivo Properties of Human DNA Methyltransferases from Double-Stranded Methylation Patterns

DNA methyltransferases establish methylation patterns in cells and transmit these patterns over cell generations, thereby influencing each cell's epigenetic states. Three primary DNA methyltransferases have been identified in mammals: DNMT1, DNMT3A and DNMT3B. Extensive in vitro studies have investigated key properties of these enzymes, namely their substrate specificity and processivity. Here we study these properties in vivo, by applying novel statistical analysis methods to double-stranded DNA methylation patterns collected using hairpin-bisulfite PCR. Our analysis fits a novel Hidden Markov Model (HMM) to the observed data, allowing for potential bisulfite conversion errors, and yields statistical estimates of parameters that quantify enzyme processivity and substrate specificity. We apply this model to methylation patterns established in vivo at three loci in humans: two densely methylated inactive X (Xi)-linked loci ( and ), and an autosomal locus (), where methylation densities are tissue-specific but moderate. We find strong evidence for a high level of processivity of DNMT1 at and , with the mean association tract length being a few hundred base pairs. Regardless of tissue types, methylation patterns at are dominated by DNMT1 maintenance events, similar to the two Xi-linked loci, but are insufficiently informative regarding processivity to draw any conclusions about processivity at that locus. At all three loci we find that DNMT1 shows a strong preference for adding methyl groups to hemi-methylated CpG sites over unmethylated sites. The data at all three loci also suggest low (possibly 0) association of the de novo methyltransferases, the DNMT3s, and are consequently uninformative about processivity or preference of these enzymes. We also extend our HMM to reanalyze published data on mouse DNMT1 activities in vitro. The results suggest shorter association tracts (and hence weaker processivity), and much longer non-association tracts than human DNMT1 in vivo

Persistent activation of interlinked type 2 airway epithelial gene networks in sputum-derived cells from aeroallergen-sensitized symptomatic asthmatics

© 2018 The Author(s). Atopic asthma is a persistent disease characterized by intermittent wheeze and progressive loss of lung function. The disease is thought to be driven primarily by chronic aeroallergen-induced type 2-associated inflammation. However, the vast majority of atopics do not develop asthma despite ongoing aeroallergen exposure, suggesting additional mechanisms operate in conjunction with type 2 immunity to drive asthma pathogenesis. We employed RNA-Seq profiling of sputum-derived cells to identify gene networks operative at baseline in house dust mite-sensitized (HDM S ) subjects with/without wheezing history that are characteristic of the ongoing asthmatic state. The expression of type 2 effectors (IL-5, IL-13) was equivalent in both cohorts of subjects. However, in HDM S -wheezers they were associated with upregulation of two coexpression modules comprising multiple type 2- and epithelial-associated genes. The first module was interlinked by the hubs EGFR, ERBB2, CDH1 and IL-13. The second module was associated with CDHR3 and mucociliary clearance genes. Our findings provide new insight into the molecular mechanisms operative at baseline in the airway mucosa in atopic asthmatics undergoing natural aeroallergen exposure, and suggest that susceptibility to asthma amongst these subjects involves complex interactions between type 2- and epithelial-associated gene networks, which are not operative in equivalently sensitized/exposed atopic non-asthmatics

The immunogenicity and pathogenicity of Pasteurella multocida isolated from poultry in Namakkal

The growth of Indian poultry industry is phenomenal and the industry now ranks 5(th) in the world in egg production. Intensification of poultry farming was the way by which this growth was obtained. Such intensification has led to health problems associated with both existing and emerging diseases. Of the diseases affecting poultry, fowl cholera is a very important bacterial disease. Fowl cholera occurs worldwide and causes economic losses due to increased mortality, increased number of culls and a marked drop in egg production. The causative agent of fowl cholera namely Pasturella multocida was first isolated in 1878 by Bollinger (Carter and Chengappa, 1986). A recent survey on the prevalence of fowl cholera in Namakkal area reported an incidence of 4.17% in 0-10 wks, 8.47% in growers and 3.61% in layers (Daniel et al., 1998). There is little information regarding the serotype or pathogenicity of the Indian Pasturella multocida isolates associated with fowl cholera outbreaks. Though commercial vaccines are being used in chicken breeding farms, occasional outbreaks of fowl cholera still occur. Hence, this study was undertaken to characterize local isolates of Pasturella multocida associated with outbreaks in Namakkal as a first step in the development of a fowl cholera vaccine