A novel nonparametric computational strategy for identifying differential methylation regions

Background DNA methylation has long been known as an epigenetic gene silencing mechanism. For a motivating example, the methylomes of cancer and non-cancer cells show a number of methylation differences, indicating that certain features characteristics of cancer cells may be related to methylation characteristics. Robust methods for detecting differentially methylated regions (DMRs) could help scientists narrow down genome regions and even find biologically important regions. Although some statistical methods were developed for detecting DMR, there is no default or strongest method. Fisher’s exact test is direct, but not suitable for data with multiple replications, while regression-based methods usually come with a large number of assumptions. More complicated methods have been proposed, but those methods are often difficult to interpret. Results In this paper, we propose a three-step nonparametric kernel smoothing method that is both flexible and straightforward to implement and interpret. The proposed method relies on local quadratic fitting to find the set of equilibrium points (points at which the first derivative is 0) and the corresponding set of confidence windows. Potential regions are further refined using biological criteria, and finally selected based on a Bonferroni adjusted t-test cutoff. Using a comparison of three senescent and three proliferating cell lines to illustrate our method, we were able to identify a total of 1077 DMRs on chromosome 21. Conclusions We proposed a completely nonparametric, statistically straightforward, and interpretable method for detecting differentially methylated regions. Compared with existing methods, the non-reliance on model assumptions and the straightforward nature of our method makes it one competitive alternative to the existing statistical methods for defining DMRs.

Gene of the month: H3F3A and H3F3B

H3F3A and H3F3B genes are located at 1q42.12 and 17q25.1, respectively, and encode identical H3.3 core histone proteins which form part of the histone hetero-octamer complex. Histones function by packaging DNA into small units, the nucleosome, and are highly susceptible to epigenetic post-translational modification. H3 K27 mutations have been shown to inhibit the polycomb repressive complex 2, which is normally involved in epigenetic gene silencing. Mutations in H3F3A and H3F3B are increasingly recognised in a variety of solid tumours. Point mutations in H3F3A have been described in giant cell tumour of bone and paediatric-type diffuse high-grade gliomas. Mutations in H3F3B have been described in chondroblastoma. Loss of trimethylation of H3 K27 is characteristic of most sporadic and radiation-associated malignant peripheral nerve sheath tumours. Immunohistochemistry with a variety of novel antibodies directed against specific mutations, as well as loss of H3K27me3 staining, may be useful in specific settings and in diagnostically challenging cases.

An enhancer screen identifies new suppressors of small-RNA-mediated epigenetic gene silencing

Small non-protein coding RNAs are involved in pathways that control the genome at the level of chromatin. In Schizosaccharomyces pombe, small interfering RNAs (siRNAs) are required for the faithful propagation of heterochromatin that is found at peri-centromeric repeats. In contrast to repetitive DNA, protein-coding genes are refractory to siRNA-mediated heterochromatin formation, unless siRNAs are expressed in mutant cells. Here we report the identification of 20 novel mutant alleles that enable de novo formation of heterochromatin at a euchromatic protein-coding gene by using trans-acting siRNAs as triggers. For example, a single amino acid substitution in the pre-mRNA cleavage factor Yth1 enables siRNAs to trigger silent chromatin formation with unparalleled efficiency. Our results are consistent with a kinetic nascent transcript processing model for the inhibition of small-RNA-directed de novo formation of heterochromatin and lay a foundation for further mechanistic dissection of cellular activities that counteract epigenetic gene silencing.

Publisher Correction: An epigenetic gene silencing pathway selectively acting on transgenic DNA in the green alga Chlamydomonasis

A Correction to this paper has been published: https://doi.org/10.1038/s41467-020-20623-0

An epigenetic gene silencing pathway selectively acting on transgenic DNA in the green alga Chlamydomonas

Silencing of exogenous DNA can make transgene expression very inefficient. Genetic screens in the model alga Chlamydomonas have demonstrated that transgene silencing can be overcome by mutations in unknown gene(s), thus producing algal strains that stably express foreign genes to high levels. Here, we show that the silencing mechanism specifically acts on transgenic DNA. Once a permissive chromatin structure has assembled, transgene expression can persist even in the absence of mutations disrupting the silencing pathway. We have identified the gene conferring the silencing and show it to encode a sirtuin-type histone deacetylase. Loss of gene function does not appreciably affect endogenous gene expression. Our data suggest that transgenic DNA is recognized and then quickly inactivated by the assembly of a repressive chromatin structure composed of deacetylated histones. We propose that this mechanism may have evolved to provide protection from potentially harmful types of environmental DNA.

Downregulation of the Polycomb-Associated Methyltransferase Ezh2 during Maturation of Hippocampal Neurons Is Mediated by MicroRNAs Let-7 and miR-124

Ezh2 is a catalytic subunit of the polycomb repressive complex 2 (PRC2) which mediates epigenetic gene silencing through depositing the mark histone H3 lysine 27 trimethylation (H3K27me3) at target genomic sequences. Previous studies have demonstrated that Enhancer of Zeste Homolog 2 (Ezh2) was differentially expressed during maturation of hippocampal neurons; in immature neurons, Ezh2 was abundantly expressed, whereas in mature neurons the expression Ezh2 was significantly reduced. Here, we report that Ezh2 is downregulated by microRNAs (miRs) that are expressed during the hippocampal maturation process. We show that, in mature hippocampal neurons, lethal-7 (let-7) and microRNA-124 (miR-124) are robustly expressed and can target cognate motifs at the 3′-UTR of the Ezh2 gene sequence to downregulate Ezh2 expression. Together, these data demonstrate that the PRC2 repressive activity during hippocampal maturation is controlled through a post-transcriptional mechanism that mediates Ezh2 downregulation in mature neurons.

The PRC2-dependent epigenetic reprograming of the bladder epithelium exacerbates urinary tract infections

Mucosal imprint sensitizes recurrent urinary tract infections (UTIs), a significant health and quality of life burden worldwide, which are associated with heightened inflammatory host response, severe basal cell hyperplasia and impaired superficial cell differentiation. Here, we show that bladder infections induce expression of Ezh2, the methyltransferase of polycomb repressor complex 2 (PRC2)-dependent epigenetic gene silencing program. In mouse models of UTIs, urothelium-specific inactivation of PRC2 reduces the urine bacteria burden. The mutants exhibit a blunted inflammatory response likely due to the diminished activity of NF-κB signaling pathway. PRC2 inactivation also improves urothelial differentiation and attenuates basal cell hyperplasia phenotype. Moreover, the Ezh2-specific small molecule inhibitors markedly improve disease outcomes of bladder superinfection and chronic cystitis. Taken together, these findings suggest that the UTI-induced epigenetic reprograming in the bladder urothelium likely contributes to the mucosal imprint, and further suggest that targeting PRC2 methyltransferase offers a non-antibiotic strategy to mitigate UTIs.