A FUNCTIONAL EPIGENETIC CLOCK FOR RATS

Abstract Evidence from humans suggests that incorporation of phenotypic aging measures in the development of epigenetic clocks leads to more functionally relevant biomarkers. As a result, the aim of this study was to utilize a deeply phenotyped rat cohort—that included data from rotarod, open field, frailty index, and FACS—to generate a novel epigenetic clock. DNA methylation was assessed via reduced representation bisulfite sequencing (RRBS) for n=142 male Fischer rats from NIA aging colony, ranging in age from 1 to 27 months. Phenotypic traits were combined to generate an multi-system aging measure that was then used to train the epigenetic clock. Using an independent validation sample, age-adjusted epigenetic clock measures were associated with numerous traits, including: open field time resting (p=0.005), open field time climbing (p=0.001), body weight (p=0.02), and rotarod max (p=0.04). In moving forward, it will be important to examine cross-species comparisons, longitudinal change, and functional enrichment.

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A rat epigenetic clock recapitulates phenotypic aging and co-localizes with heterochromatin

eLife ◽

10.7554/elife.59201 ◽

2020 ◽

Vol 9 ◽

Author(s):

Morgan Levine ◽

Ross A McDevitt ◽

Margarita Meer ◽

Kathy Perdue ◽

Andrea Di Francesco ◽

...

Keyword(s):

Epigenetic Clock ◽

Reduced Representation ◽

Fischer 344 ◽

Biomarkers Of Aging ◽

Cell Counts ◽

Reduced Representation Bisulfite Sequencing ◽

Epigenetic Age ◽

Intergenic Regions ◽

F344 Rats ◽

Physiological And Biochemical

Robust biomarkers of aging have been developed from DNA methylation in humans and more recently, in mice. This study aimed to generate a novel epigenetic clock in rats—a model with unique physical, physiological, and biochemical advantages—by incorporating behavioral data, unsupervised machine learning, and network analysis to identify epigenetic signals that not only track with age, but also relates to phenotypic aging. Reduced representation bisulfite sequencing (RRBS) data was used to train an epigenetic age (DNAmAge) measure in Fischer 344 CDF (F344) rats. This measure correlated with age at (r = 0.93) in an independent sample, and related to physical functioning (p=5.9e-3), after adjusting for age and cell counts. DNAmAge was also found to correlate with age in male C57BL/6 mice (r = 0.79), and was decreased in response to caloric restriction. Our signatures driven by CpGs in intergenic regions that showed substantial overlap with H3K9me3, H3K27me3, and E2F1 transcriptional factor binding.

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Multi-Omics Characterization of Inflammatory Bowel Disease-Induced Hyperplasia/Dysplasia in the Rag2−/−/Il10−/− Mouse Model

International Journal of Molecular Sciences ◽

10.3390/ijms22010364 ◽

2020 ◽

Vol 22 (1) ◽

pp. 364

Author(s):

Qiyuan Han ◽

Thomas J. Y. Kono ◽

Charles G. Knutson ◽

Nicola M. Parry ◽

Christopher L. Seiler ◽

...

Keyword(s):

Inflammatory Bowel Disease ◽

Bowel Disease ◽

Gastrointestinal Disease ◽

Tumor Development ◽

Proximal Colon ◽

Helicobacter Hepaticus ◽

Reduced Representation ◽

Reduced Representation Bisulfite Sequencing ◽

Inflammatory Bowel

Epigenetic dysregulation is hypothesized to play a role in the observed association between inflammatory bowel disease (IBD) and colon tumor development. In the present work, DNA methylome, hydroxymethylome, and transcriptome analyses were conducted in proximal colon tissues harvested from the Helicobacter hepaticus (H. hepaticus)-infected murine model of IBD. Reduced representation bisulfite sequencing (RRBS) and oxidative RRBS (oxRRBS) analyses identified 1606 differentially methylated regions (DMR) and 3011 differentially hydroxymethylated regions (DhMR). These DMR/DhMR overlapped with genes that are associated with gastrointestinal disease, inflammatory disease, and cancer. RNA-seq revealed pronounced expression changes of a number of genes associated with inflammation and cancer. Several genes including Duox2, Tgm2, Cdhr5, and Hk2 exhibited changes in both DNA methylation/hydroxymethylation and gene expression levels. Overall, our results suggest that chronic inflammation triggers changes in methylation and hydroxymethylation patterns in the genome, altering the expression of key tumorigenesis genes and potentially contributing to the initiation of colorectal cancer.

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Methylation Marks of Blood Leukocytes of Native Hucul Mares Differentiated in Age

International Journal of Genomics ◽

10.1155/2019/2839614 ◽

2019 ◽

Vol 2019 ◽

pp. 1-9

Author(s):

T. Ząbek ◽

E. Semik-Gurgul ◽

T. Szmatoła ◽

A. Gurgul ◽

A. Fornal ◽

...

Keyword(s):

Cancer Progression ◽

Late Onset ◽

Advanced Age ◽

Farm Animals ◽

Blood Leukocytes ◽

Data Set ◽

Reduced Representation ◽

Cellular Signal

Horses are one of the longest-living species of farm animals. Advanced age is often associated with a decrease in body condition, dysfunction of immune system, and late-onset disorders. Due to this, the search for new solutions in the prevention and treatment of pathological conditions of the advanced age of horses is desirable. That is why the identification of aging-related changes in the horse genome is interesting in this respect. In the recent years, the research on aging includes studies of age-related epigenetic effects observed on the DNA methylation level. We applied reduced representation bisulfite sequencing (RRBS) to uncover a range of age DMR sites in genomes of blood leukocytes derived from juvenile and aged horses of native Hucul breed. Genes colocated with age-related differentially methylated regions (age DMRs) are the members of pathways involved in cellular signal transduction, immune response, neurogenesis, differentiation, development, and cancer progression. A positive correlation was found between methylation states and gene expression in particular loci from our data set. Some of described age DMR-linked genes were also reported elsewhere. Obtained results contribute to the knowledge about the molecular basis of aging of equine blood cells.

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Epigenetic Changes in the Regulation of Nicotiana tabacum Response to Cucumber Mosaic Virus Infection and Symptom Recovery through Single-Base Resolution Methylomes

Viruses ◽

10.3390/v10080402 ◽

2018 ◽

Vol 10 (8) ◽

pp. 402 ◽

Cited By ~ 10

Author(s):

Chenguang Wang ◽

Chaonan Wang ◽

Wenjie Xu ◽

Jingze Zou ◽

Yanhong Qiu ◽

...

Keyword(s):

Immune Response ◽

Nicotiana Tabacum ◽

Viral Infection ◽

Cucumber Mosaic Virus ◽

Mosaic Virus ◽

Developmental Stages ◽

Reduced Representation ◽

Reduced Representation Bisulfite Sequencing ◽

Symptom Recovery

Plants have evolved multiple mechanisms to respond to viral infection. These responses have been studied in detail at the level of host immune response and antiviral RNA silencing (RNAi). However, the possibility of epigenetic reprogramming has not been thoroughly investigated. Here, we identified the role of DNA methylation during viral infection and performed reduced representation bisulfite sequencing (RRBS) on tissues of Cucumber mosaic virus (CMV)-infected Nicotiana tabacum at various developmental stages. Differential methylated regions are enriched with CHH sequence contexts, 80% of which are located on the gene body to regulate gene expression in a temporal style. The methylated genes depressed by methyltransferase inhibition largely overlapped with methylated genes in response to viral invasion. Activation in the argonaute protein and depression in methyl donor synthase revealed the important role of dynamic methylation changes in modulating viral clearance and resistance signaling. Methylation-expression relationships were found to be required for the immune response and cellular components are necessary for the proper defense response to infection and symptom recovery.

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Technical Considerations for Reduced Representation Bisulfite Sequencing with Multiplexed Libraries

Journal of Biomedicine and Biotechnology ◽

10.1155/2012/741542 ◽

2012 ◽

Vol 2012 ◽

pp. 1-8 ◽

Cited By ~ 38

Author(s):

Aniruddha Chatterjee ◽

Euan J. Rodger ◽

Peter A. Stockwell ◽

Robert J. Weeks ◽

Ian M. Morison

Keyword(s):

Bisulfite Sequencing ◽

Flow Cell ◽

Throughput Capacity ◽

Sequencing Data ◽

Bioinformatics Pipeline ◽

Illumina Hiseq ◽

Reduced Representation ◽

Reduced Representation Bisulfite Sequencing ◽

Unique Mapping ◽

Nucleotide Resolution

Reduced representation bisulfite sequencing (RRBS), which couples bisulfite conversion and next generation sequencing, is an innovative method that specifically enriches genomic regions with a high density of potential methylation sites and enables investigation of DNA methylation at single-nucleotide resolution. Recent advances in the Illumina DNA sample preparation protocol and sequencing technology have vastly improved sequencing throughput capacity. Although the new Illumina technology is now widely used, the unique challenges associated with multiplexed RRBS libraries on this platform have not been previously described. We have made modifications to the RRBS library preparation protocol to sequence multiplexed libraries on a single flow cell lane of the Illumina HiSeq 2000. Furthermore, our analysis incorporates a bioinformatics pipeline specifically designed to process bisulfite-converted sequencing reads and evaluate the output and quality of the sequencing data generated from the multiplexed libraries. We obtained an average of 42 million paired-end reads per sample for each flow-cell lane, with a high unique mapping efficiency to the reference human genome. Here we provide a roadmap of modifications, strategies, and trouble shooting approaches we implemented to optimize sequencing of multiplexed libraries on an a RRBS background.

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Characterization of Global DNA Methylation in Different Gene Regions Reveals Candidate Biomarkers in Pigs with High and Low Levels of Boar Taint

Veterinary Sciences ◽

10.3390/vetsci7020077 ◽

2020 ◽

Vol 7 (2) ◽

pp. 77 ◽

Cited By ~ 1

Author(s):

Xiao Wang ◽

Haja N. Kadarmideen

Keyword(s):

Dna Methylation ◽

Cpg Island ◽

Cpg Islands ◽

Gene Interaction ◽

Boar Taint ◽

Reduced Representation ◽

Gene Interaction Networks ◽

Reduced Representation Bisulfite Sequencing ◽

Pathways Analysis ◽

Porcine Gene

DNA methylation of different gene components, including different exons and introns, or different lengths of exons and introns is associated with differences in gene expression. To investigate the methylation of porcine gene components associated with the boar taint (BT) trait, this study used reduced representation bisulfite sequencing (RRBS) data from nine porcine testis samples in three BT groups (low, medium and high BT). The results showed that the methylation levels of the first exons and first introns were lower than those of the other exons and introns. The first exons/introns of CpG island regions had even lower levels of methylation. A total of 123 differentially methylated promoters (DMPs), 194 differentially methylated exons (DMEs) and 402 differentially methylated introns (DMIs) were identified, of which 80 DMPs (DMP-CpGis), 112 DMEs (DME-CpGis) and 166 DMIs (DMI-CpGis) were discovered in CpG islands. Importantly, GPX1 contained one each of DMP, DME, DMI, DMP-CpGi, DME-CpGi and DMI-CpGi. Gene-GO term relationships and pathways analysis showed DMP-CpGi-related genes are mainly involved in methylation-related biological functions. In addition, gene–gene interaction networks consisted of nodes that were hypo-methylated GPX1, hypo-methylated APP, hypo-methylated ATOX1, hyper-methylated ADRB2, hyper-methylated RPS6KA1 and hyper-methylated PNMT. They could be used as candidate biomarkers for reducing boar taint in pigs, after further validation in large cohorts.

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