Distinct Methylation Patterns Research Articles

Evolutionary Contribution of Duplicated Genes to Genome Evolution in the Ginseng Species Complex.

Genes duplicated by whole genome duplication (WGD) and small-scale duplication (SSD) have played important roles in adaptive evolution of all flowering plants. However, it still remains underinvestigated how the distinct models of duplication events and their contending evolutionary patterns have shaped the genome and epigenomes of extant plant species. In this study, we investigated the contribution of the WGD- and SSD-derived duplicate genes to the genome evolution of one diploid and three closely related allotetraploid Panax species based on genome, methylome, and proteome data sets. Our genome-wide comparative analyses revealed that although the ginseng species complex was recently diverged, they have evolved distinct overall patterns of nucleotide variation, cytosine methylation, and protein-level expression. In particular, genetic and epigenetic asymmetries observed in the recent WGD-derived genes are largely consistent across the ginseng species complex. In addition, our results revealed that gene duplicates generated by ancient WGD and SSD mechanisms exhibited distinct evolutionary patterns. We found the ancient WGD-derived genes (i.e., ancient collinear gene) are genetically more conserved and hypomethylated at the cytosine sites. In contrast, some of the SSD-derived genes (i.e., dispersal duplicated gene) showed hypermethylation and high variance in nucleotide variation pattern. Functional enrichment analyses of the duplicated genes indicated that adaptation-related traits (i.e., photosynthesis) created during the distant ancient WGDs are further strengthened by both the more recent WGD and SSD. Together, our findings suggest that different types of duplicated genes may have played distinct but relaying evolutionary roles in the polyploidization and speciation processes in the ginseng species complex.

DNA methylation-based signatures classify sporadic pituitary tumors according to clinicopathological features.

Distinct genome-wide methylation patterns cluster pituitary neuroendocrine tumors (PitNETs) into molecular groups associated with specific clinicopathological features. Here we aim to identify, characterize, and validate methylation signatures that objectively classify PitNET into clinicopathological groups. Combining in-house and publicly available data, we conducted an analysis of the methylome profile of a comprehensive cohort of 177 tumors (Panpit cohort) and 20 nontumor specimens from the pituitary gland. We also retrieved methylome data from an independent PitNET cohort (N = 86) to validate our findings. We identified three methylation clusters associated with adenohypophyseal cell lineages and functional status using an unsupervised approach. Differentially methylated probes (DMP) significantly distinguished the Panpit clusters and accurately assigned the samples of the validation cohort to their corresponding lineage and functional subtypes memberships. The DMPs were annotated in regulatory regions enriched with enhancer elements, associated with pathways and genes involved in pituitary cell identity, function, tumorigenesis, and invasiveness. Some DMPs correlated with genes with prognostic and therapeutic values in other intra- or extracranial tumors. We identified and validated methylation signatures, mainly annotated in enhancer regions that distinguished PitNETs by distinct adenohypophyseal cell lineages and functional status. These signatures provide the groundwork to develop an unbiased approach to classifying PitNETs according to the most recent classification recommended by the 2017 WHO and to explore their biological and clinical relevance in these tumors.

Distinct DNA Methylation Patterns of Rheumatoid Arthritis Peripheral Blood and Synovial Tissue T Cells

ObjectiveTo study epigenetic patterns in T lymphocytes that accumulate in the rheumatoid arthritis (RA) synovium, we characterized DNA methylation of CD3+ T cells in peripheral blood and synovial tissue in patients with RA and osteoarthritis (OA).MethodsGenomic DNA of CD3+ T cells was isolated from patients with RA (n = 8) and OA (n = 5) from blood or the synovium at the time of an arthroplasty using antibodies and magnetic beads. Methylation was measured by using the Illumina Infinium MethylationEPIC Kit. Differentially methylated loci (DML) and differentially methylated genes (DMGs) were identified by using Welch’s t‐test. Principal component analysis, hierarchical clustering, and pathway analysis were used to determine relationships among groups.ResultsWhen we compared DNA methylation of CD3+ T cells between peripheral blood and synovial tissue within each disease, 4615 and 164 DML were identified in RA and OA samples, respectively, resulting in 832 and 36 DMGs. A principal component analysis showed that methylation differences in T cells were greater on the basis of on location (blood vs synovium) rather than disease (RA vs OA). Differentially modified pathways were significantly enriched between RA blood and synovial T cells, especially in genes related to complement, integrin cell surface interactions, and the P53 pathway. The limited number of DMGs identified between OA blood and synovial T cells did not conform to biologic pathways.ConclusionThe patterns of DNA methylation in RA show location‐specific differences related to immune pathways, whereas methylation differences in OA are limited. The RA joint‐specific signatures could be due to selective accumulation of T‐cell populations or expansion of differentially marked adaptive immune cells. Understanding epigenetic patterns could provide clues to the types of T cells that accumulate in the RA joint and identify potential therapeutic targets.

Identification of methylation changes associated with positive and negative growth deviance in Gambian infants using a targeted methyl sequencing approach of genomic DNA.

Low birthweight and reduced height gain during infancy (stunting) may arise at least in part from adverse early life environments that trigger epigenetic reprogramming that may favor survival. We examined differential DNA methylation patterns using targeted methyl sequencing of regions regulating gene activity in groups of rural Gambian infants: (a) low and high birthweight (DNA from cord blood (n = 16 and n = 20, respectively), from placental trophoblast tissue (n = 21 and n = 20, respectively), and DNA from peripheral blood collected from infants at 12 months of age (n = 23 and n = 17, respectively)), and, (b) the top 10% showing rapid postnatal length gain (high, n = 20) and the bottom 10% showing slow postnatal length gain (low, n = 20) based on z score change between birth and 12 months of age (LAZ) (DNA from peripheral blood collected from infants at 12 months of age). Using BiSeq analysis to identify significant methylation marks, for birthweight, four differentially methylated regions (DMRs) were identified in trophoblast DNA, compared to 68 DMRs in cord blood DNA, and 54 DMRs in 12‐month peripheral blood DNA. Twenty‐five DMRs were observed to be associated with high and low length for age (LAZ) at 12 months. With the exception of five loci (associated with two different genes), there was no overlap between these groups of methylation marks. Of the 194 CpG methylation marks contained within DMRs, 106 were located to defined gene regulatory elements (promoters, CTCF‐binding sites, transcription factor‐binding sites, and enhancers), 58 to gene bodies (introns or exons), and 30 to intergenic DNA. Distinct methylation patterns associated with birthweight between comparison groups were observed in DNA collected at birth (at the end of intrauterine growth window) compared to those established by 12 months (near the infancy/childhood growth transition). The longitudinal differences in methylation patterns may arise from methylation adjustments, changes in cellular composition of blood or both that continue during the critical postnatal growth period, and in response to early nutritional and infectious environmental exposures with impacts on growth and longer‐term health outcomes.

DNA methylation patterns identify subgroups of pancreatic neuroendocrine tumors with clinical association

Here we report the DNA methylation profile of 84 sporadic pancreatic neuroendocrine tumors (PanNETs) with associated clinical and genomic information. We identified three subgroups of PanNETs, termed T1, T2 and T3, with distinct patterns of methylation. The T1 subgroup was enriched for functional tumors and ATRX, DAXX and MEN1 wild-type genotypes. The T2 subgroup contained tumors with mutations in ATRX, DAXX and MEN1 and recurrent patterns of chromosomal losses in half of the genome with no association between regions with recurrent loss and methylation levels. T2 tumors were larger and had lower methylation in the MGMT gene body, which showed positive correlation with gene expression. The T3 subgroup harboured mutations in MEN1 with recurrent loss of chromosome 11, was enriched for grade G1 tumors and showed histological parameters associated with better prognosis. Our results suggest a role for methylation in both driving tumorigenesis and potentially stratifying prognosis in PanNETs.

Dynamic Changes of Fetal-Derived Hypermethylated RASSF1A and Septin 9 Sequences in Maternal Plasma.

DNA methylation has a tissue-specific feature, and placenta has distinct methylation patterns from peripheral blood cells. Although fetal/placental-derived cell free DNA (cfDNA) in the maternal blood has been reported in recent decades, systematic exploration of dynamic changes of the placental epigenetic signatures across gestation is lacking. The primary goal of this study was to characterize prenatal and postnatal methylation levels of placental-sourced RASSF1A and Septin 9 sequences in maternal plasma. Here, we used a quantitative methylation-sensitive PCR (qMS-PCR) assay to check the methylation status of RASSF1A and Septin 9 in placental tissues of pregnant women and plasma samples from non-pregnant individuals. Then, we examined the methylation levels of the two targets in maternal plasma from expectant women at different gestational ages and postdelivery. Hypermethylated RASSF1A and Septin 9 were identified in placental samples but undetectable in peripheral blood of healthy non-pregnant women. Further, hypermethylated RASSF1A sequence was found in all three trimesters of pregnancy except for early gestation (8weeks). Moreover, methylation scores of the two targets increased as pregnancy progressed. In addition, hypermethylated RASSF1A sequence was detectable in maternal plasma from 12h (one case) to 24h postdelivery (three cases) in 18 pregnant women. Our data on the variation of fetal-sourced methylated RASSF1A levels in maternal plasma in relation to gestational age provide a useful basis for improving the reliability of the methylation assay for non-invasive prenatal diagnosis (NIPD) in clinical practice.

Epigenetic Modifications in Acute Lymphoblastic Leukemia: From Cellular Mechanisms to Therapeutics.

Epigenetic modification pattern is considered as a characteristic feature in blood malignancies. Modifications in the DNA methylation modulators are recurrent in lymphoma and leukemia, so that the distinct methylation pattern defines different types of leukemia. Generally, the role of epigenetics is less understood, and most investigations are focused on genetic abnormalities and cytogenic studies to develop novel treatments for patients with hematologic disorders. Recently, understanding the underlying mechanism of acute lymphoblastic leukemia (ALL), especially epigenetic alterations as a driving force in the development of ALL opens a new era of investigation for developing promising strategy, beyond available conventional therapy. This review will focus on a better understanding of the epigenetic mechanisms in cancer development and progression, with an emphasis on epigenetic alterations in ALL including, DNA methylation, histone modification, and microRNA alterations. Other topics that will be discussed include the use of epigenetic alterations as a promising therapeutic target in order to develop novel, well-suited approaches against ALL. According to the literature review, leukemogenesis of ALL is extensively influenced by epigenetic modifications, particularly DNA hyper-methylation, histone modification, and miRNA alteration.

DNA methylation profiling identifies epigenetic differences between early versus late stages of diabetic chronic kidney disease.

We investigated a cross-sectional epigenome-wide association study of patients with early and late diabetes-associated chronic kidney disease (CKD) to identify possible epigenetic differences between the two groups as well as changes in methylation across all stages of diabetic CKD. We also evaluated the potential of using a panel of identified 5'-C-phosphate-G-3' (CpG) sites from this cohort to predict the progression of diabetic CKD. This cross-sectional study recruited 119 adults. DNA was extracted from blood using the Qiagen QIAampDNA Mini Spin Kit. Genome-wide methylation analysis was performed using Illumina Infinium MethylationEPIC BeadChips (HM850K). Intensity data files were processed and analysed using the minfi and MissMethyl packages for R. We examined the degree of methylation of CpG sites in early versus late diabetic CKD patients for CpG sites with an unadjusted P-value <0.01 and an absolute change in methylation of 5% (n = 239 CpG sites). Hierarchical clustering of the 239 CpG sites largely separated the two groups. A heat map for all 239 CpG sites demonstrated distinct methylation patterns in the early versus late groups, with CpG sites showing evidence of progressive change. Based on our differentially methylated region (DMR) analysis of the 239 CpG sites, we highlighted two DMRs, namely the cysteine-rich secretory protein 2 (CRISP2) and piwi-like RNA-mediated gene silencing 1(PIWIL1) genes. The best predictability for the two groups involved a receiver operating characteristics curve of eight CpG sites alone and achieved an area under the curve of 0.976. We have identified distinct DNA methylation patterns between early and late diabetic CKD patients as well as demonstrated novel findings of potential progressive methylation changes across all stages (1-5) of diabetic CKD at specific CpG sites. We have also identified associated genes CRISP2 and PIWIL1, which may have the potential to act as stage-specific diabetes-associated CKD markers, and showed that the use of a panel of eight identified CpG sites alone helps to increase the predictability for the two groups.

Systematic integrated analysis of genetic and epigenetic variation in diabetic kidney disease

Poor metabolic control and host genetic predisposition are critical for diabetic kidney disease (DKD) development. The epigenome integrates information from sequence variations and metabolic alterations. Here, we performed a genome-wide methylome association analysis in 500 subjects with DKD from the Chronic Renal Insufficiency Cohort for DKD phenotypes, including glycemic control, albuminuria, kidney function, and kidney function decline. We show distinct methylation patterns associated with each phenotype. We define methylation variations that are associated with underlying nucleotide variations (methylation quantitative trait loci) and show that underlying genetic variations are important drivers of methylation changes. We implemented Bayesian multitrait colocalization analysis (moloc) and summary data-based Mendelian randomization to systematically annotate genomic regions that show association with kidney function, methylation, and gene expression. We prioritized 40 loci, where methylation and gene-expression changes likely mediate the genotype effect on kidney disease development. Functional annotation suggested the role of inflammation, specifically, apoptotic cell clearance and complement activation in kidney disease development. Our study defines methylation changes associated with DKD phenotypes, the key role of underlying genetic variations driving methylation variations, and prioritizes methylome and gene-expression changes that likely mediate the genotype effect on kidney disease pathogenesis.

Pathogenic Differences of Type 1 Restriction-Modification Allele Variants in Experimental Listeria monocytogenes Meningitis.

Background: L. monocytogenes meningoencephalitis has a mortality rate of up to 50% and neurofunctional sequelae are common. Type I restriction-modification systems (RMS) are capable of adding methyl groups to the host genome. Some contain multiple sequence recognition (hsdS) genes that recombine, resulting in distinct DNA methylation patterns and patterns of gene expression. These phenotypic switches have been linked to virulence and have recently been discovered in multiple clonal complexes of L. monocytogenes. In the present study, we investigated the significant of RMS on L. monocytogenes virulence during the acute phase of experimental meningitis.Methods: L. monocytogenes strains containing RMS systems were identified, and purified clones enriched for single hsdS alleles were isolated. In vivo, 11-day old Wistar rats were infected with an inoculum containing (a) one of 4 single RMS allele variants (A, B, C, D) treated with amoxicillin (AMX 50 mg/kg/dosis, q8h), (b) a mixture of all 4 variants with or without AMX treatment, or (c) different mixtures of 2 RMS allele variants. At selected time points after infection, clinical and inflammatory parameters, bacterial titers and brain damage were determined. Changes in the relative frequency of the occurring RMS alleles in the inoculum and in CSF or cerebellum of infected animals were analyzed by capillary electrophoresis.Results: We have identified a phase variable RMS locus within L. monocytogenes CC4 and generated stocks that stably expressed each of the possible hsdS genes within that loci. Generation of these allele variants (A, B, C, D) allowed us to determine the methylation pattern associated with each hsdS through SMRT sequencing. In vivo infections with these single allele variants revealed differences in disease severity in that C induced the worst clinical outcome and more pronounced hippocampal apoptosis; D showed the most pronounced weight loss and the highest bacterial titer in the cerebellum. A caused the least severe disease.Conclusion: We identified that L. monocytogenes expressing hsdS (A) causes less damage than when other hsdS genes are expressed. While expression of hsdSC and D worsened the outcome in L. monocytogenes meningitis. We also demonstrate a competitive advantage of variants C and B over variant A in this model. Phenotypical switching may therefore represent a mechanism of virulence regulation during the acute phase of CNS infections with L. monocytogenes.

Genome-wide DNA methylation analysis using MethylCap-seq in canine high-grade B-cell lymphoma.

DNA methylation is a comprehensively studied epigenetic modification and plays crucial roles in cancer development. In the present study, MethylCap-seq was used to characterize the genome-wide DNA methylation patterns in canine high-grade B-cell lymphoma (cHGBL). Canine methylated DNA fragments were captured and the MEDIUM-HIGH and LOW fraction of methylated DNA was obtained based on variation in CpG methylation density. In the MEDIUM-HIGH and LOW fraction, 2144 and 1987 cHGBL-specific hypermethylated genes, respectively, were identified. Functional analysis highlighted pathways strongly related to oncogenesis. The relevant signaling pathways associated with neuronal system were also revealed, echoing recent novel findings that neurogenesis plays key roles in tumor establishment. In addition, 14 genes were hypermethylated in all the cHGBL cases but not in the healthy dogs. These genes might be potential signatures for tracing cHGBL, and some of them have been reported to play roles in various types of cancers. Further, the distinct methylation pattern of cHGBL showed a concordance with the clinical outcome, suggesting that aberrant epigenetic changes may influence tumor behavior. In summary, our study characterized genome-wide DNA methylation patterns using MethylCap-seq in cHGBL; the findings suggest that specific DNA hypermethylation holds promise for dissecting tumorigenesis and uncovering biomarkers for monitoring the progression of cHGBL.

Epigenetic stratification of head and neck cancer survivors reveals differences in lycopene levels, alcohol consumption, and methylation of immune regulatory genes

BackgroundInflammation has been associated with higher rates of recurrence and mortality in head and neck cancer (HNC). While the biological mechanisms predisposing patients to heightened inflammatory states remain largely unknown, DNA methylation has been proposed to reflect systemic inflammation. In this analysis, we attempt to identify meaningful epigenetic patterns in HNC survivors by stratifying individuals based on DNA methylation profiles in leukocytes.ResultsWe used hierarchical clustering to uncover three distinct methylation patterns among HNC survivors. Each group displayed a unique methylation signature in inflammatory pathways including cytokine and B-cell receptor signaling. Additionally, we examined physiological, clinical, and lifestyle parameters related to inflammation, such as circulating carotenoid and cytokine levels, cancer treatment type, and alcohol consumption. Specifically, we identified one group of survivors who had significant differential methylation of transcriptional and translational regulators as well as genes in the T-cell receptor signaling pathway, including hypermethylation of CD40 ligand (CD40LG) and Tec protein tyrosine kinase (TEC) and hypomethylation of CD8A. This group also displayed high circulating lycopene levels. We identified another group that had distinctive methylation in the toll-like receptor (TLR) signaling pathway, including hypomethylation of TLR5, a component of the inhibitor of nuclear factor-kappa B kinase complex (CHUK), and two mitogen-activated protein kinases (MAP3K8 and MAP2K3). This group also had hypermethylation of mitochondrial ribosomal genes along with higher rates of alcohol consumption.ConclusionThe correlation between lycopene, alcohol consumption, DNA methylation, and inflammation warrants further investigation and may have implications in future recommendations and interventions to impact health outcomes in HNC survivors.

Distinct promoter methylation patterns of LKB1 in the hamartomatous polyps of Peutz-Jeghers syndrome and its potential in gastrointestinal malignancy prediction

BackgroundPeutz-Jeghers Syndrome (PJS) is known as a rare inherited polyposis due to the malfunction of serine/threonine kinase gene LKB1. However, not all of PJS patients carry LKB1 germline mutation. Previous researches have observed the elevated DNA methylation level in PJS polyps. Nevertheless, the mechanism of such abnormal and its impact on PJS patients remains to be fully described.ResultsThe results proved a significant increase on the methylation level of LKB1 promoter in PJS polyps compared with normal colon biopsies through bisulfite PCR followed by Sanger sequencing. Moreover, the methylation pattern in PJS polyps could be further categorized as three different scenarios: hypermethylated, hemimethylated and hypomethylated pattern. Furthermore, immunohistochemistry of DNMT1/3a/3b suggested the up-regulation of DNMT1 and 3a might participate the epigenetic alternation of LKB1 in PJS polyps. Logistic regression suggested hypomethylated LKB1 promoter in PJS polyps as a risk factor for gastrointestinal malignancies in PJS patients.ConclusionsThe promoter methylation level of LKB1 gene in PJS polyps is generally elevated compared with normal colon mucosa. Yet not all of PJS polyps carry hypermethylated LKB1 promoter. Hypomethylation in this region has linked to malignant tumors in PJS patients. Given the rarity of PJS, this work together with previous researches, have proved the importance of LKB1 promoter methylation in PJS development and prognosis.

Discovery of Molecular DNA Methylation-Based Biomarkers through Genome-Wide Analysis of Response Patterns to BCG for Bladder Cancer.

Background: Bacillus Calmette-Guérin (BCG) immunotherapy, the standard adjuvant intravesical therapy for some intermediate and most high-risk non-muscle invasive bladder cancers (NMIBCs), suffers from a heterogenous response rate. Molecular markers to help guide responses are scarce and currently not used in the clinical setting. Methods: To identify novel biomarkers and pathways involved in response to BCG immunotherapy, we performed a genome-wide DNA methylation analysis of NMIBCs before BCG therapy. Genome-wide DNA methylation profiles of DNA isolated from tumors of 26 BCG responders and 27 failures were obtained using the Infinium MethylationEPIC BeadChip. Results: Distinct DNA methylation patterns were found by genome-wide analysis in the two groups. Differentially methylated CpG sites were predominantly located in gene promoters and gene bodies associated with bacterial invasion of epithelial cells, chemokine signaling, endocytosis, and focal adhesion. In total, 40 genomic regions with a significant difference in methylation between responders and failures were detected. The differential methylation state of six of these regions, localized in the promoters of the genes GPR158, KLF8, C12orf42, WDR44, FLT1, and CHST11, were internally validated by bisulfite-sequencing. GPR158 promoter hypermethylation was the best predictor of BCG failure with an AUC of 0.809 (p-value < 0.001). Conclusions: Tumors from BCG responders and BCG failures harbor distinct DNA methylation profiles. Differentially methylated DNA regions were detected in genes related to pathways involved in bacterial invasion of cells or focal adhesion. We identified candidate DNA methylation biomarkers that may help to predict patient prognosis after external validation in larger, well-designed cohorts.

The integrated genomic and epigenomic landscape of brainstem glioma

Brainstem gliomas are a heterogeneous group of tumors that encompass both benign tumors cured with surgical resection and highly lethal cancers with no efficacious therapies. We perform a comprehensive study incorporating epigenetic and genomic analyses on a large cohort of brainstem gliomas, including Diffuse Intrinsic Pontine Gliomas. Here we report, from DNA methylation data, distinct clusters termed H3-Pons, H3-Medulla, IDH, and PA-like, each associated with unique genomic and clinical profiles. The majority of tumors within H3-Pons and-H3-Medulla harbors H3F3A mutations but shows distinct methylation patterns that correlate with anatomical localization within the pons or medulla, respectively. Clinical data show significantly different overall survival between these clusters, and pathway analysis demonstrates different oncogenic mechanisms in these samples. Our findings indicate that the integration of genetic and epigenetic data can facilitate better understanding of brainstem gliomagenesis and classification, and guide future studies for the development of novel treatments for this disease.

What Have We Learned from GWAS for Atopic Dermatitis?

GWASs have revealed multiple loci associated with atopic dermatitis (AD). Some have confirmed pre-existing knowledge, including the role of skin barrier and type 2 inflammation in AD pathogenesis, whereas others have provided newer insights, including evidence of autoimmunity and previously unrecognized genes controlling epidermal differentiation. The majority of risk loci are in intergenic regions for which functional mechanism(s) remain unknown. These loci require detailed molecular studies carried out in cells and tissues of relevance to AD. Genomic findings to date account for ∼30% of AD heritability, therefore, considerable further work is needed to fully understand individual risk.

Silencing of Mutator Elements in Maize Involves Distinct Populations of Small RNAs and Distinct Patterns of DNA Methylation.

Transposable elements (TEs) are a ubiquitous feature of plant genomes. Because of the threat they post to genome integrity, most TEs are epigenetically silenced. However, even closely related plant species often have dramatically different populations of TEs, suggesting periodic rounds of activity and silencing. Here, we show that the process of de novo methylation of an active element in maize involves two distinct pathways, one of which is directly implicated in causing epigenetic silencing and one of which is the result of that silencing. Epigenetic changes involve changes in gene expression that can be heritably transmitted to daughter cells in the absence of changes in DNA sequence. Epigenetics has been implicated in phenomena as diverse as development, stress response, and carcinogenesis. A significant challenge facing those interested in investigating epigenetic phenomena is determining causal relationships between DNA methylation, specific classes of small RNAs, and associated changes in gene expression. Because they are the primary targets of epigenetic silencing in plants and, when active, are often targeted for de novo silencing, TEs represent a valuable source of information about these relationships. We use a naturally occurring system in which a single TE can be heritably silenced by a single derivative of that TE. By using this system it is possible to unravel causal relationships between different size classes of small RNAs, patterns of DNA methylation, and heritable silencing. Here, we show that the long terminal inverted repeats within Zea mays MuDR transposons are targeted by distinct classes of small RNAs during epigenetic silencing that are dependent on distinct silencing pathways, only one of which is associated with transcriptional silencing of the transposon. Further, these small RNAs target distinct regions of the terminal inverted repeats, resulting in different patterns of cytosine methylation with different functional consequences with respect to epigenetic silencing and the heritability of that silencing.

Characterization of histone modification patterns and prediction of novel promoters using functional principal component analysis.

Characterization of distinct histone methylation and acetylation binding patterns in promoters and prediction of novel regulatory regions remains an important area of genomic research, as it is hypothesized that distinct chromatin signatures may specify unique genomic functions. However, methods that have been proposed in the literature are either descriptive in nature or are fully parametric and hence more restrictive in pattern discovery. In this article, we propose a two-step non-parametric statistical inference procedure to characterize unique histone modification patterns and apply it to analyzing the binding patterns of four histone marks, H3K4me2, H3K4me3, H3K9ac, and H4K20me1, in human B-lymphoblastoid cells. In the first step, we used a functional principal component analysis method to represent the concatenated binding patterns of these four histone marks around the transcription start sites as smooth curves. In the second step, we clustered these curves to reveal several unique classes of binding patterns. These uncovered patterns were used in turn to scan the whole-genome to predict novel and alternative promoters. Our analyses show that there are three distinct promoter binding patterns of active genes. Further, 19654 regions not within known gene promoters were found to overlap with human ESTs, CpG islands, or common SNPs, indicative of their potential role in gene regulation, including being potential novel promoter regions.

Comprehensive analyses of 723 transcriptomes enhance genetic and biological interpretations for complex traits in cattle.

By uniformly analyzing 723 RNA-seq data from 91 tissues and cell types, we built a comprehensive gene atlas and studied tissue specificity of genes in cattle. We demonstrated that tissue-specific genes significantly reflected the tissue-relevant biology, showing distinct promoter methylation and evolution patterns (e.g., brain-specific genes evolve slowest, whereas testis-specific genes evolve fastest). Through integrative analyses of those tissue-specific genes with large-scale genome-wide association studies, we detected relevant tissues/cell types and candidate genes for 45 economically important traits in cattle, including blood/immune system (e.g., CCDC88C) for male fertility, brain (e.g., TRIM46 and RAB6A) for milk production, and multiple growth-related tissues (e.g., FGF6 and CCND2) for body conformation. We validated these findings by using epigenomic data across major somatic tissues and sperm. Collectively, our findings provided novel insights into the genetic and biological mechanisms underlying complex traits in cattle, and our transcriptome atlas can serve as a primary source for biological interpretation, functional validation, studies of adaptive evolution, and genomic improvement in livestock.

Distinct Expression and Methylation Patterns for Genes with Different Fates following a Single Whole-Genome Duplication in Flowering Plants.

For most sequenced flowering plants, multiple whole-genome duplications (WGDs) are found. Duplicated genes following WGD often have different fates that can quickly disappear again, be retained for long(er) periods, or subsequently undergo small-scale duplications. However, how different expression, epigenetic regulation, and functional constraints are associated with these different gene fates following a WGD still requires further investigation due to successive WGDs in angiosperms complicating the gene trajectories. In this study, we investigate lotus (Nelumbo nucifera), an angiosperm with a single WGD during the K–pg boundary. Based on improved intraspecific-synteny identification by a chromosome-level assembly, transcriptome, and bisulfite sequencing, we explore not only the fundamental distinctions in genomic features, expression, and methylation patterns of genes with different fates after a WGD but also the factors that shape post-WGD expression divergence and expression bias between duplicates. We found that after a WGD genes that returned to single copies show the highest levels and breadth of expression, gene body methylation, and intron numbers, whereas the long-retained duplicates exhibit the highest degrees of protein–protein interactions and protein lengths and the lowest methylation in gene flanking regions. For those long-retained duplicate pairs, the degree of expression divergence correlates with their sequence divergence, degree in protein–protein interactions, and expression level, whereas their biases in expression level reflecting subgenome dominance are associated with the bias of subgenome fractionation. Overall, our study on the paleopolyploid nature of lotus highlights the impact of different functional constraints on gene fate and duplicate divergence following a single WGD in plant.