The DNA Methylation Maintenance Protein UHRF1 Regulates Fetal Globin Expression Independent of HBG Promoter DNA Methylation

Abstract Pharmacological or genetic induction of fetal hemoglobin (HbF, α2γ2) in adult red blood cells is a proven strategy to ameliorate the clinical symptoms of sickle cell disease (SCD) and β-thalassemia. Therefore, efforts are underway to better understand mechanisms that mediate the perinatal switch from HbF to adult hemoglobin (HbA, α2β2). We performed a CRISPR-Cas9/guide (g) RNA screen to identify novel proteins that regulate HbF production in HUDEP-2 cells, a human erythroid line that normally expresses HbA. We identified UHRF1 (ubiquitin-like with PHD and RING finger domains 1) as a repressor of HbF production. UHRF1 binds hemi-methylated DNA and recruit DNA methyltransferase 1 (DNMT1) to ensure faithful maintenance of DNA methylation during DNA replication. Numerous UHRF1-interacting proteins, including DNMT1, EHMT1/2 and HDAC2 are associated with γ-globin repression. We used CRISPR/Cas9 and RNA interference to validate UHRF1 as a HbF regulator. Compared to non-targeting gRNA UHRF1 disruption using Cas9 + 2 separate gRNAs increased the γ-globin/γ+β-globin RNA ratio from 1.9 to 25.8/27.1% (P<0.01), increased the fraction of HbF immunostaining cells ("F-cells") from 7.5 to 25.1/35.4% and increased HbF protein from 2.10 to 16.3/15.0% (P<0.01) in HUDEP-2 cells. Compared to a control luciferase shRNA, 2 different UHRF1 shRNAs increased theγ-globin/γ+β-globin RNA ratio from 9.68% to 21.59/28.93% (P<0.01), increased the F-cell fraction from 37.9 to 49.8/55.6% and increased HbF protein from 9.1 to 16.18/18.5% (P<0.05) in erythroid cells derived from normal adult peripheral blood CD34+ cells. UHRF1 deficiency did not alter erythroid maturation or expression of key transcription factor genes that regulate HbF expression in HUDEP-2 or CD34+ cells (BCL11A, ZBTB7A, MYB and KLF1). UHRF1 mutant proteins defective in recognizing H3K9me2 (FW237/238AA), binding to hemi-methylated DNA (R491A) or ubiquitination of H3K23 to enhance DNMT1 recruitment (C741A), were unable to repress HBG1/HBG2. These mutations have the most profound effects on maintaining DNA methylation, indicating that UHRF1 represses HBG1/HBG2 in HUDEP-2 cells through this mechanism. UHRF1 knockout induced genome-wide demethylation including 6 CpG sites located at positions -162, -53, -50, +6, +17, +50 positions relative to the γ-globin (HBG1 and HBG2) transcription start site. Demethylation of these sites is thought to be required for γ-globin de-repression. However, forced demethylation of these cytosines in HUDEP-2 cells using specific gRNAs + dead (d) Cas9-TET1 was not sufficient to activate γ-globin expression when UHRF1 was present. Additionally, dCas9-DNMT3a-mediated methylation of the HBG promoter CpG residues in UHRF1 knockdown HUDEP-2 cells did not inhibit γ-globin expression in UHRF1 knockout HUDEP-2 cells. Based on these studies, we conclude that: 1) UHRF1 regulates γ-globin transcription; 2) demethylation of CpG sites at the HBG gene promoters is neither necessary or sufficient for γ-globin induction; 3) UHRF1 regulates γ-to-β globin gene switching either by methylating DNA regions other than those present around the HBG promoter or through non-canonical activities. Distinguishing these mechanisms will elucidate further our understanding of globin gene switching and could identify new pathways for pharmacological induction of HbF. Disclosures No relevant conflicts of interest to declare.

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A natural DNMT1 mutation elevates the fetal hemoglobin via epigenetic de-repression of γ-globin gene in β-thalassemia

Blood ◽

10.1182/blood.2020006425 ◽

2020 ◽

Author(s):

Yi Gong ◽

Xinhua Zhang ◽

Qianqian Zhang ◽

Yanxia Zhang ◽

Yuhua Ye ◽

...

Keyword(s):

Dna Methylation ◽

Dna Methyltransferase ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Dna Methyltransferase 1 ◽

Macromolecular Complexes ◽

Cell Pattern ◽

Cd34 Cells ◽

Epigenetic Regulator ◽

Mutant Cells

DNA methyltransferase 1 (DNMT1) is a major epigenetic regulator of the formation of large macromolecular complexes that repress human γ-globin expression by maintaining DNA methylation. However, very little is known about the association of DNMT1 variants with β-thalassemia phenotypes. We systematically investigated associations between variants in DNMT1 and phenotypes in 1,142 β-thalassemia subjects and identified a novel missense mutation (c.2633G>A, S878F) in the bromo-adjacent homology-1 (BAH1) domain of DNMT1. We functionally characterized this mutation in CD34+ cells from patientsand engineered HuDEP-2 mutant cells. Our results demonstrate that DNMT1 phosphorylation is abrogated by substitution of serine with phenylalanine at position 878, resulting in lower stability and loss of catalytic activity. S878F mutation also attenuated the interactions of DNMT1 with BCL11A, GATA1 and HDAC1/2 and reduced the recruitment of DNMT1 to the HBG promoters, leading to epigenetic de-repression of γ-globin expression. By analyzing the F-cell pattern, we demonstrated that the effect of DNMT1 mutation on increased fetal hemoglobin (HbF) is heterocellular. Furthermore, introduction of S878F mutation into erythroid cells by CRISPR-Cas9 recapitulated γ-globin reactivation. Thus, the natural S878F DNMT1 mutation is a novel modulator of HbF synthesis and represents a potential new therapeutic target for β-hemoglobinopathies.

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Abstract 40: Disturbed Flow Alters Genomewide DNA Methylation Patterns, Regulating Endothelial Gene Expression and Atherosclerosis

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.34.suppl_1.40 ◽

2014 ◽

Vol 34 (suppl_1) ◽

Author(s):

Jessilyn Dunn ◽

Haiwei Qiu ◽

Soyeon Kim ◽

Daudi Jjingo ◽

Ryan Hoffman ◽

...

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Dna Methyltransferase ◽

Methylation Status ◽

Western Diet ◽

Dependent Manner ◽

Gene Promoters ◽

Genome Wide ◽

Methylation Patterns ◽

Endothelial Gene Expression

Atherosclerosis preferentially occurs in arterial regions of disturbed blood flow (d-flow), which alters gene expression, endothelial function, and atherosclerosis. Here, we show that d-flow regulates genome-wide DNA methylation patterns in a DNA methyltransferase (DNMT)-dependent manner. We found that d-flow induced expression of DNMT1, but not DNMT3a or DNMT3b, in mouse arterial endothelium in vivo and in cultured endothelial cells by oscillatory shear (OS) compared to unidirectional laminar shear in vitro. The DNMT inhibitor 5-Aza-2’deoxycytidine (5Aza) or DNMT1 siRNA significantly reduced OS-induced endothelial inflammation. Moreover, 5Aza reduced lesion formation in two atherosclerosis models using ApoE-/- mice (western diet for 3 months and the partial carotid ligation model with western diet for 3 weeks). To identify the 5Aza mechanisms, we conducted two genome-wide studies: reduced representation bisulfite sequencing (RRBS) and transcript microarray using endothelial-enriched gDNA and RNA, respectively, obtained from the partially-ligated left common carotid artery (LCA exposed to d-flow) and the right contralateral control (RCA exposed to s-flow) of mice treated with 5Aza or vehicle. D-flow induced DNA hypermethylation in 421 gene promoters, which was significantly prevented by 5Aza in 335 genes. Systems biological analyses using the RRBS and the transcriptome data revealed 11 mechanosensitive genes whose promoters were hypermethylated by d-flow but rescued by 5Aza treatment. Of those, five genes contain hypermethylated cAMP-response-elements in their promoters, including the transcription factors HoxA5 and Klf3. Their methylation status could serve as a mechanosensitive master switch in endothelial gene expression. Our results demonstrate that d-flow controls epigenomic DNA methylation patterns in a DNMT-dependent manner, which in turn alters endothelial gene expression and induces atherosclerosis.

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Structural insights into CpG-specific DNA methylation by human DNA methyltransferase 3B

Nucleic Acids Research ◽

10.1093/nar/gkaa111 ◽

2020 ◽

Vol 48 (7) ◽

pp. 3949-3961 ◽

Cited By ~ 5

Author(s):

Chien-Chu Lin ◽

Yi-Ping Chen ◽

Wei-Zen Yang ◽

James C K Shen ◽

Hanna S Yuan

Keyword(s):

Dna Methylation ◽

Dna Methyltransferase ◽

Cytosine Methylation ◽

Catalytic Domain ◽

De Novo ◽

Water Channel ◽

Dna Methyltransferases ◽

Structural Basis ◽

Cpg Sites ◽

Methylation Patterns

Abstract DNA methyltransferases are primary enzymes for cytosine methylation at CpG sites of epigenetic gene regulation in mammals. De novo methyltransferases DNMT3A and DNMT3B create DNA methylation patterns during development, but how they differentially implement genomic DNA methylation patterns is poorly understood. Here, we report crystal structures of the catalytic domain of human DNMT3B–3L complex, noncovalently bound with and without DNA of different sequences. Human DNMT3B uses two flexible loops to enclose DNA and employs its catalytic loop to flip out the cytosine base. As opposed to DNMT3A, DNMT3B specifically recognizes DNA with CpGpG sites via residues Asn779 and Lys777 in its more stable and well-ordered target recognition domain loop to facilitate processive methylation of tandemly repeated CpG sites. We also identify a proton wire water channel for the final deprotonation step, revealing the complete working mechanism for cytosine methylation by DNMT3B and providing the structural basis for DNMT3B mutation-induced hypomethylation in immunodeficiency, centromere instability and facial anomalies syndrome.

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Complex DNA sequence readout mechanisms of the DNMT3B DNA methyltransferase

Nucleic Acids Research ◽

10.1093/nar/gkaa938 ◽

2020 ◽

Vol 48 (20) ◽

pp. 11495-11509

Author(s):

Michael Dukatz ◽

Sabrina Adam ◽

Mahamaya Biswal ◽

Jikui Song ◽

Pavel Bashtrykov ◽

...

Keyword(s):

Dna Methylation ◽

Dna Methyltransferase ◽

Catalytic Mechanism ◽

Catalytic Domain ◽

Dna Methyltransferases ◽

Flanking Sequence ◽

Cpg Sites ◽

Binding Interface ◽

Target Sites ◽

Flanking Sequences

Abstract DNA methyltransferases interact with their CpG target sites in the context of variable flanking sequences. We investigated DNA methylation by the human DNMT3B catalytic domain using substrate pools containing CpX target sites in randomized flanking context and identified combined effects of CpG recognition and flanking sequence interaction together with complex contact networks involved in balancing the interaction with different flanking sites. DNA methylation rates were more affected by flanking sequences at non-CpG than at CpG sites. We show that T775 has an essential dynamic role in the catalytic mechanism of DNMT3B. Moreover, we identify six amino acid residues in the DNA-binding interface of DNMT3B (N652, N656, N658, K777, N779, and R823), which are involved in the equalization of methylation rates of CpG sites in favored and disfavored sequence contexts by forming compensatory interactions to the flanking residues including a CpG specific contact to an A at the +1 flanking site. Non-CpG flanking preferences of DNMT3B are highly correlated with non-CpG methylation patterns in human cells. Comparison of the flanking sequence preferences of human and mouse DNMT3B revealed subtle differences suggesting a co-evolution of flanking sequence preferences and cellular DNMT targets.

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Sensitivity of Diffuse Large B-Cell Lymphomas to DNA Methyltransferase Inhibitors Is Associated with a Specific Epigenetic Signature.

Blood ◽

10.1182/blood.v108.11.831.831 ◽

2006 ◽

Vol 108 (11) ◽

pp. 831-831

Author(s):

Megan Ryan ◽

Leandro Cerchietti ◽

Maria E. Figueroa ◽

John Greally ◽

Ari Melnick

Keyword(s):

Dna Methylation ◽

B Cell ◽

Cell Lines ◽

Dna Methyltransferase ◽

Serum Starvation ◽

Gene Promoters ◽

B Cell Lymphomas ◽

Response Curves ◽

Differentially Methylated Genes ◽

Large B Cell

Abstract DNA methyltransferase inhibitor drugs (MTIs) such as decitabine can overcome gene silencing due to aberrant hypermethylation of gene promoters. Presumably, this effect is responsible for the therapeutic activity of MTIs as clinically demonstrated in myelodysplasias (MDS) and leukemias. Other tumors such as diffuse large B-cell lymphomas (DLBCLs) can also present with aberrant promoter hypermethylation. However, it is currently difficult to prospectively identify patients likely to respond to MTIs, since specific methylation markers or signatures have not yet been identified. We predicted that decitabine would have anti-lymphoma activity in a subset of DLBCLs, and that these cases would exhibit specific methylation signatures predictive of response to these drugs. To determine whether this is the case we first exposed a panel of 7 DLBCL cell lines (Ly1, Ly7, Ly10, SU-DHL6, Farage, Pfeiffer and Toledo) to increasing concentrations of decitabine (0.5, 1, 2.5, 5, 10, 50 and 100 μM) administered after synchronization by 12 hr serum starvation. Viability was assessed after 48 hr of culture by MTS-based assay and Trypan blue exclusion. The IC25 and IC50 were calculated for all cell lines by constructing dose-response curves. The IC25 was used to discriminate sensitive (6.3 ± 1.2 μM) vs. resistant (49.4 ± 5 μM, p < 0.01) cell lines. Interestingly, there was no correlation between MTI sensitivity and DLBCL subtype as defined by recent gene expression profiling classification efforts (i.e. GCB vs. ABC, or BCR vs. OxPhos). To identify the methylation signatures of these DLBCL cells we used a method that we developed for genome-wide DNA methylation quantification called HELP (HpaII tiny fragment Enrichment by LM-PCR). HELP is based on comparative Msp1 and HpaII digestion of genomic DNA, followed by size specific amplification and co-hybridization to custom high-density oligonucleotide arrays designed to provide uniform data collection over 25,000 promoters. HELP compares favorably to other high throughput methods in that it is highly reproducible (R > 0.98) and has an extremely robust signal-to-noise ratio. DNA was collected from the DLBCL cells for HELP prior to drug treatment. Most significantly we found that unsupervised (i.e. unbiased) clustering of DNA methylation profiles could readily segregate decitabine resistant vs. sensitive DLBCL cell lines. Correspondence analysis clearly identified a methylation signature consisting of 133 differentially methylated genes that distinguishes between decitabine sensitive and resistant cells. Most of these appeared to be functionally relevant including such genes as Caspase-9, RARB, JUNB, and ELK1. Biological assays to determine the contribution of these genes to the phenotype are underway. Taken together, our data suggest that MTIs might be effective in a cohort of DLBCL cases that exhibit the specific methylation signature that we have identified. Prospective evaluation of the predictive value of this signature may allow optimal selection of patients for clinical trials with these agents.

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DNA Methylation Changes Following 5-Azacitidine Treatment in Patients with Myelodysplastic Syndrome.

Blood ◽

10.1182/blood.v114.22.2791.2791 ◽

2009 ◽

Vol 114 (22) ◽

pp. 2791-2791

Author(s):

Huong Thi Thanh Tran ◽

Hee Nam Kim ◽

Yeo-Kyeoung Kim ◽

Jae-Sook Ahn ◽

Il-Kwon Lee ◽

...

Keyword(s):

Dna Methylation ◽

Tumor Suppressor ◽

Tumor Suppressor Genes ◽

Dna Methyltransferase ◽

Conflicts Of Interest ◽

Control Group ◽

Disease Pathogenesis ◽

Suppressor Genes ◽

Clinical Responses ◽

Dependent Probe

Abstract Abstract 2791 Poster Board II-767 Gene silencing by promoter methylation is as potent as functional inactivating of tumor suppressor genes by mutations. DNA methyltransferase inhibitor, 5-azacytidine (AC) and 5-aza-2 -deoxycitidine (DAC), which is proved to be effective in myelodysplastic syndromes (MDS) can induce re-expression in cancer; however their mechanism remains controversial. 25 tumor suppressor genes by MS-MLPA (methylation-specific multiplex ligation-dependent probe amplification) were analyzed in 44 MDS patients treated Vidaza® (5-azacitidine, AC). Hypermethylation of at least one gene was detected in 9/44 patients (20.5%), including four genes CDKN2B, FHIT, ESR1 and IGSF4. Interestingly, of 9 hypermethylated patients, 8 patients showed demethylation in concordance with their clinical responses after three to five cycles AC treatment. Lack or decrease methylation was observed in four patients with hematological improvements. Persistence methylation was observed in four others who became AML transformation or no response after treatment, especially reinforcing methylated gene in a case progressed to leukemia later. Our study also founds out IGSF4 gene hypermethylation in MDS as a first report. Additionally, mRNA expression of CDKN2B, IGSF4, and ESR1 in MDS were significantly lower than those in the control group (p < 0.05). Our results suggest that the methylation changes of specific genes contributes to disease pathogenesis and might present a molecular marker that can be used to monitor the efficacy of AC treatment in MDS. Disclosures: No relevant conflicts of interest to declare.

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Inhibition of LSD1 Influences Multiple Mechanisms of Epigenetic Gene Regulation During Terminal Erythroid Maturation

Blood ◽

10.1182/blood.v120.21.3442.3442 ◽

2012 ◽

Vol 120 (21) ◽

pp. 3442-3442

Author(s):

Michael Getman ◽

Samantha J England ◽

James Palis ◽

Laurie A Steiner

Keyword(s):

Dna Methylation ◽

Dna Methyltransferase ◽

Molecular Mechanisms ◽

Conflicts Of Interest ◽

Histone Demethylase ◽

Enzyme Linked Immunosorbent Assay ◽

Local Effects ◽

Erythroid Progenitor ◽

Significant Difference ◽

Erythroid Maturation

Abstract Abstract 3442 The maturation of a committed erythroid progenitor to a functional red blood cell is a complex process involving significant changes in gene expression during a time of rapid cell division and nuclear condensation. LSD1 (Lysine-Specific Histone Demethylase 1) is a histone H3 lysine 4 (H3K4) and lysine 9 (H3K9) demethylase that plays pivotal role in this process. LSD1 participates in both enhancer and repressor complexes, and facilitates repression of γ-globin by participating in the Direct Repeat Erythroid Complex (Cui, MCB, 2011). LSD1 inhibitors Tranylcypromine (TCP) and Pargyline (PG) are being investigated as potential therapies for the β-globinopathies, however little is known about the broader functional or genomic consequences of LSD1 inhibition on terminal erythroid maturation. Both TCP and PG impair erythroid maturation in Extensively Self Renewing Erythroblasts (ESREs), a primary cell model of terminal erythroid maturation. ESREs are primary cells derived from fetal liver that proliferate extensively in culture, but retain the ability to appropriately mature and enucleate (England, Blood, 2011), making them ideal for functional and genomic studies of terminal erythroid maturation. In untreated or vehicle (DMSO) treated cultures >90% of cells are benzidine positive by day3 of maturation. In contrast, cultures treated with 400um PG, 1um TCP, or 2um TCP were 72, 42, and 33% benzidine positive by maturation day3, respectively. Cells in the TCP-and PG- treated cultures also had morphologic evidence of impaired maturation, with larger nuclei and more basophilic cytoplasm. In addition to its role as a histone demethylase, LSD1 stabilizes DNMT1 (DNA methyltransferase 1; Wang, Nat Genet 2009). We hypothesized that loss of DNA methylation contributes to the maturation impairment seen with LSD1 inhibitors, and that inhibition of DNMTs with decitabine would also impair terminal erythroid maturation. Consistent with this hypothesis, ESREs treated with decitabine demonstrated a dose-dependent impairment of maturation similar to that seen with PG and TCP. To elucidate the molecular mechanisms underlying the maturation impairment in TCP- and PG- treated cultures, levels of H3K4me2 and methylated DNA (5-methyl cytosine, 5-mC) were assessed both globally and at specific loci. An ELISA (Enzyme-linked Immunosorbent Assay) was used to assess global levels of H3K4me2 and 5-mC in vehicle-, PG-, and TCP-treated cultures after 24 hours of maturation. Global levels of H3K4me2 were significantly higher in PG- and TCP- treated samples than control. In maturing cells, there was no significant difference in the level of 5-mC in vehicle- and inhibitor- treated cultures. It is well established, however, that global DNA methylation decreases with erythroid maturation (Seashore, Science, 2011), and a significant decrease in 5-mC occurs in ESREs during the first 24hrs of maturation. As TCP- and vehicle- treated cultures mature differently, the effect of TCP on 5-mC levels was also assessed in self-renewing ESREs at the proerythroblast stage. Unlike maturing cells, TCP-treated proerythroblasts had a significant decrease in 5-mC levels compared to control. Chromatin immunoprecipitation (ChIP) was used to examine the local effects of LSD1 inhibition on H3K4me2 enrichment at erythroid-specific promoters. TCP-treated cultures had non-uniform changes in H3K4me2 enrichment, with levels increased at some promoters (e.g. protein 4.1,εy-globin), but unchanged at others (e.g. β-globin). To further study the relationship between LSD1 inhibition and H3K4me2 levels, ChIP-seq was used to identify LSD1 sites that co-localized with putative enhancers, defined as peaks of H3K4me2 binding > than 1kb from a transcription start site. ChIP-qPCR was used to compare the level of H3K4me2 at 5 validated enhancer-associated LSD1 sites in vehicle- and TCP-treated cells. The effect of TCP was variable, with only 2/5 enhancer-associated LSD1 sites having increased H3K4me2. Lastly, the local effects of inhibitors on 5-mC were examined using a methyl binding domain pulldown coupled with qPCR. In TCP-treated cells, 5-mC levels declined at several loci, most notably at the εy-globin promoter. Taken together, these results suggest that the impaired erythroid maturation associated with LSD1 inhibition results from the perturbation of multiple mechanisms of epigenetic regulation. Disclosures: No relevant conflicts of interest to declare.

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Naturally occurring methylation inhibitor: DNA hypomethylation and hemoglobin synthesis in human K562 cells.

Molecular and Cellular Biology ◽

10.1128/mcb.7.5.1759 ◽

1987 ◽

Vol 7 (5) ◽

pp. 1759-1763 ◽

Cited By ~ 14

Author(s):

S B Lyon ◽

L Buonocore ◽

M Miller

Keyword(s):

Dna Methylation ◽

Genomic Dna ◽

Dna Methyltransferase ◽

Globin Gene ◽

K562 Cells ◽

Gene Probe ◽

Dna Hypomethylation ◽

Naturally Occurring ◽

Methylation Inhibitor

A naturally occurring methylation inhibitor isolated from rabbit liver and named methinin inhibits a number of methyltransferases. Methinin is a low-molecular-weight compound (1,400) that has an active amine group. This compound inhibits the DNA methyltransferase of human erythroleukemia cells (K562) in vitro. When the K562 cells were grown in medium containing methinin, fetal hemoglobin was produced. Small but detectable amounts of adult hemoglobin were also produced. Methinin was not toxic to these cells. The overall rate of genomic DNA methylation was reduced by 60% in cells grown in medium containing methinin. Southern blots of genomic DNA from methinin-treated cells and untreated cells hybridized to a 32P-labeled globin gene probe showed that one site in the globin gene region was hypomethylated. Methinin is a naturally occurring compound which inhibits DNA methylation both in vitro and in vivo.

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Differential binding of EKLF to embryonic/fetal globin gene promoters during development: Model for globin gene switching

Blood Cells Molecules and Diseases ◽

10.1016/j.bcmd.2006.10.139 ◽

2007 ◽

Vol 38 (2) ◽

pp. 181-182

Author(s):

Tim M. Townes ◽

Dewang Zhou ◽

Kevin M. Pawlik ◽

Jinxiang Ren ◽

Chiao-Wang Sun

Keyword(s):

Globin Gene ◽

Development Model ◽

Gene Promoters ◽

Differential Binding ◽

Gene Switching

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A genomic predictor of lifespan in vertebrates

Scientific Reports ◽

10.1038/s41598-019-54447-w ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 7

Author(s):

Benjamin Mayne ◽

Oliver Berry ◽

Campbell Davies ◽

Jessica Farley ◽

Simon Jarman

Keyword(s):

Dna Methylation ◽

Gene Promoters ◽

Epigenetic Changes ◽

Ecological Significance ◽

Biological Functions ◽

Maximum Lifespan ◽

Genome Sequences ◽

Biological Ageing ◽

Cpg Sites ◽

Whole Genomes

AbstractBiological ageing and its mechanistic underpinnings are of immense biomedical and ecological significance. Ageing involves the decline of diverse biological functions and places a limit on a species’ maximum lifespan. Ageing is associated with epigenetic changes involving DNA methylation. Furthermore, an analysis of mammals showed that the density of CpG sites in gene promoters, which are targets for DNA methylation, is correlated with lifespan. Using 252 whole genomes and databases of animal age and promotor sequences, we show a pattern across vertebrates. We also derive a predictive lifespan clock based on CpG density in a selected set of promoters. The lifespan clock accurately predicts maximum lifespan in vertebrates (R2 = 0.76) from the density of CpG sites within only 42 selected promoters. Our lifespan clock provides a wholly new method for accurately estimating lifespan using genome sequences alone and enables estimation of this challenging parameter for both poorly understood and extinct species.

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