Conservation of aging and cancer epigenetic signatures across human and mouse

Aging and cancer are two interrelated processes, with aging being a major risk factor for the development of cancer. Parallel epigenetic alterations have been described for both, although differences, especially within the DNA hypomethylation scenario, have also been recently reported. While many of these observations arise from the use of mouse models, there is a lack of systematic comparisons of human and mouse epigenetic patterns in the context of disease. However, such comparisons are significant as they allow to establish the extent to which some of the observed similarities or differences arise from pre-existing species-specific epigenetic traits. Here, we have used reduced representation bisulfite sequencing to profile the brain methylomes of young and old, tumoral and non-tumoral brain samples from human and mouse. We first characterized the baseline epigenomic patterns of the species and subsequently focused on the DNA methylation alterations associated with cancer and aging. Next, we described the functional genomic and epigenomic context associated with the alterations, and finally we integrated our data to study interspecies DNA methylation levels at orthologous CpG sites. Globally, we found considerable differences between the characteristics of DNA methylation alterations in cancer and aging in both species. Moreover, we describe robust evidence for the conservation of the specific cancer and aging epigenomic signatures in human and mouse. Our observations point towards the preservation of the functional consequences of these alterations at multiple levels of genomic regulation. Finally, our analyses reveal a role for the genomic context in explaining disease- and species-specific epigenetic traits.© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution

Enhanced detection of viral RNA species using fokI-assisted digestion of DNA duplexes and DNA/RNA hybrids

The accurate detection of nucleic acids from certain biological pathogens is critical for the diagnosis of human diseases. However, amplified detection of RNA molecules from a complex sample by direct detection of RNA/DNA hybrids remains a challenge. Here, we show that type IIS endonuclease FokI is able to digest DNA duplexes and DNA/RNA hybrids when assisted by a dumbbell-like fluorescent sensing oligonucleotide. As proof of concept, we designed a battery of sensing oligonucleotides against specific regions of the SARS-CoV-2 genome and interrogated the role of FokI relaxation as a potential nicking enzyme for fluorescence signal amplification. FokI-assisted digestion of SARS-CoV-2 probes increases the detection signal of ssDNA and RNA molecules and decreases the limit of detection more than 3.5-fold as compared to conventional molecular beacon approaches. This cleavage reaction is highly specific to its target molecules, and no detection of other highly related B-coronaviruses was observed in the presence of complex RNA mixtures. In addition, the FokI-assisted reaction has a high multiplexing potential, as the combined detection of different viral RNAs, including different SARS-CoV-2 variants, was achieved in the presence of multiple combinations of fluorophores and sensing oligonucleotides. When combined with isothermal rolling circle amplification technologies, FokI-assisted digestion reduced the detection time of SARS-CoV-2 in COVID-19-positive human samples with adequate sensitivity and specificity compared to conventional reverse transcription polymerase chain reaction approaches, highlighting the potential of FokI-assisted signal amplification as a valuable sensing mechanism for the detection of human pathogens.Funding was provided by the ISCIII (COV00624 to J.R.T. andM.F.F., PI18/01527 and PI21/01067 to M.F.F.), CSIC (202020E092 to M.F.F), the European Commission NextGenerationEU, through CSIC’s Global Health Platform (PTI Salud Global) and the Spanish Ministry of Science and Innovation through the Recovery, Transformation and Resilience Plan (GL2021-03-39 and GL2021-03-040), the PCTI from the Asturias Government, co-funded by 2018−2022/FEDER (IDI/2018/146 to M.F.F.), the AECC (PROYE18061FERN to M.F.F), ISPA-Janssen (048-Intramural Nov-Tevar to J.R.T.) and the IUOPA. J.R.T is supported by a JdC fellowship from the Spanish Ministry of Science and Innovation (IJC2018-36825-I). R.F.P. and P.S.O. are supported by the Severo Ochoa program (BP17-114 and BP17-165). A.P. is supported by the PFIS program (ISCIII, FI19/ 00085). J.J.A.L. is supported by the AECC fellowship. C.M. and V.L. are supported by IUOPA, and R.G.U. is supported by CIBERER.Peer reviewe

Integrative methylome-transcriptome analysis unravels cancer cell vulnerabilities in infant MLL-rearranged B cell acute lymphoblastic leukemia

B cell acute lymphoblastic leukemia (B-ALL) is the most common childhood cancer. As predicted by its prenatal origin, infant B-ALL (iB-ALL) shows an exceptionally silent DNA mutational landscape, suggesting that alternative epigenetic mechanisms may substantially contribute to its leukemogenesis. Here, we have integrated genome-wide DNA methylome and transcriptome data from 69 patients with de novo MLL-rearranged leukemia (MLLr) and non-MLLr iB-ALL leukemia uniformly treated according to the Interfant-99/06 protocol. iB-ALL methylome signatures display a plethora of common and specific alterations associated with chromatin states related to enhancer and transcriptional control in normal hematopoietic cells. DNA methylation, gene expression, and gene coexpression network analyses segregated MLLr away from non-MLLr iB-ALL and identified a coordinated and enriched expression of the AP-1 complex members FOS and JUN and RUNX factors in MLLr iB-ALL, consistent with the significant enrichment of hypomethylated CpGs in these genes. Integrative methylome-transcriptome analysis identified consistent cancer cell vulnerabilities, revealed a robust iB-ALL–specific gene expression–correlating dmCpG signature, and confirmed an epigenetic control of AP-1 and RUNX members in reshaping the molecular network of MLLr iB-ALL. Finally, pharmacological inhibition or functional ablation of AP-1 dramatically impaired MLLr-leukemic growth in vitro and in vivo using MLLr-iB-ALL patient–derived xenografts, providing rationale for new therapeutic avenues in MLLr-iB-ALL.We thank CERCA/Generalitat de Catalunya (SGR180) and Fundació Josep Carreras-Obra Social la Caixa for their institutional support. Financial support for this work was obtained from the European Research Council (CoG-2014-646903 and PoC-2018-811220 to PM), the Spanish Ministry of Economy and Competitiveness (SAF-2019-108160-R and SAF2016-76758-R to PM and IV, respectively), the Spanish Association against cancer (AECC-CI-2015 and PROYE18061FERN to CB and MFF), the Fundación Uno entre Cienmil (to PM), the Health Institute Carlos III (ISCIII/FEDER, PI17/01028, PI15/00892, PI18/01527 to CB and AFF/MFF, respectively). We also acknowledge the Plan de Ciencia, Tecnología e Innovación from the Asturias Government cofunding 2018–2022/FEDER (IDI/2018/146to MFF). MFF also acknowledges funding from Fundación General CSIC (0348_CIE_6_E). PM also acknowledges financial support from Fundación Leo Messi. JRT and MV are supported by Juan de la Cierva fellowships by the Spanish Ministry of Science and Innovation (FJCI-2015-26965, IJC2018-36825-I, IJCI-2017-3317) and IUOPA-ISPA-FINBA (The IUOPA is supported by the Obra Social Cajastur-Liberbank, Spain). RTR is supported by a fellowship from the AECC scientific foundation. RFP and PSO are supported by the Severo Ochoa program (BP17-114 and BP17-165, respectively).Peer reviewe

Epigenetic deregulation of the histone methyltransferase KMT5B contributes to malignant transformation in glioblastoma

Glioblastoma multiforme (GBM) is the most common and aggressive type of brain tumor in adulthood. Epigenetic mechanisms are known to play a key role in GBM although the involvement of histone methyltransferase KMT5B and its mark H4K20me2 has remained largely unexplored. The present study shows that DNA hypermethylation and loss of DNA hydroxymethylation is associated with KMT5B downregulation and genome-wide reduction of H4K20me2 levels in a set of human GBM samples and cell lines as compared with non-tumoral specimens. Ectopic overexpression of KMT5B induced tumor suppressor-like features in vitro and in a mouse tumor xenograft model, as well as changes in the expression of several glioblastoma-related genes. H4K20me2 enrichment was found immediately upstream of the promoter regions of a subset of deregulated genes, thus suggesting a possible role for KMT5B in GBM through the epigenetic modulation of key target cancer genes.This research was funded by the Health Institute Carlos III (Plan Nacional de I+D+I) cofounding FEDER (PI15/00892 and PI18/01527 to MF and AF); the Government of the Principality of Asturias PCTI-Plan de Ciencia, Tecnología e Innovación de Asturias co-funding 2018–2022/FEDER (IDI/2018/146 to MF); AECC (PROYE18061FERN to MF); FGCSIC (0348_CIE_6_E to MF); Severo Ochoa Program BP17-165 to PS-O and BP17-114 to RP); the Ministry of Economy and Competitiveness of Spain (VL, Juan de la Cierva fellowship IJCI-2015-23316; JT, Juan de la Cierva fellowship FJCI-2015-26965); FICYT (AC and MG); FINBA-ISPA (VL); and IUOPA (VL and CM). The IUOPA is supported by the Obra Social Cajastur-Liberbank, Spain.Peer reviewe

Epigenetics, aging and age-related disorders

Resumen del trabajo presentado al 51st European Society of Human Genetics Conference, celebrado en Milan (Italia) del 16 al 19 de junio de 2018

Epigenome-wide analysis reveals specific DNA hypermethylation of T cells during human hematopoietic differentiation

Supplementary figures and supplementary data tables from MethylationEPIC arrays (Tejedor et al), plus a compiled dataset including raw methylation intensities, processed β-values and Phenotypic data from HumanMethylation450 arrays collected from datasets GSE63409, GSE49618, GSE88824 and GSE49031, and HumanMethylationEPIC arrays deposited in E-MTAB-6315 entry from ArrayExpress.Epigenetic regulation plays an important role in cellular development and differentiation. A detailed map of the DNA methylation dynamics that occur during cell differentiation would contribute to decipher the molecular networks governing cell fate commitment. In this study we used the most recent Illumina MethylationEPIC Beadchip platform to describe the genome-wide DNA methylation changes observed throughout hematopoietic maturation by analyzing multiple hematopoietic cell types at different developmental stages.Grants: European Commission: INFANTLEUKEMIA - GENOMIC, CELLULAR AND DEVELOPMENTAL RECONSTRUCTION OFINFANT MLL-AF4+ ACUTE LYMPHOBLASTIC LEUKEMIA (646903).Peer reviewe

Conservation of Aging and Cancer Epigenetic Signatures across Human and Mouse

Aging and cancer are two interrelated processes, with aging being a major risk factor for the development of cancer. Parallel epigenetic alterations have been described for both, although differences, especially within the DNA hypomethylation scenario, have also been recently reported. Although many of these observations arise from the use of mouse models, there is a lack of systematic comparisons of human and mouse epigenetic patterns in the context of disease. However, such comparisons are significant as they allow to establish the extent to which some of the observed similarities or differences arise from pre-existing species-specific epigenetic traits. Here, we have used reduced representation bisulfite sequencing to profile the brain methylomes of young and old, tumoral and nontumoral brain samples from human and mouse. We first characterized the baseline epigenomic patterns of the species and subsequently focused on the DNA methylation alterations associated with cancer and aging. Next, we described the functional genomic and epigenomic context associated with the alterations, and finally, we integrated our data to study interspecies DNA methylation levels at orthologous CpG sites. Globally, we found considerable differences between the characteristics of DNA methylation alterations in cancer and aging in both species. Moreover, we describe robust evidence for the conservation of the specific cancer and aging epigenomic signatures in human and mouse. Our observations point toward the preservation of the functional consequences of these alterations at multiple levels of genomic regulation. Finally, our analyses reveal a role for the genomic context in explaining disease- and species-specific epigenetic traits

Nicotinamide N-methyltransferase (NNMT) regulates the glucocorticoid signaling pathway during the early phase of adipogenesis

Abstract Obesity is associated with adipose tissue dysfunction through the differentiation and expansion of pre-adipocytes to adipocytes (hyperplasia) and/or increases in size of pre-existing adipocytes (hypertrophy). A cascade of transcriptional events coordinates the differentiation of pre-adipocytes into fully differentiated adipocytes; the process of adipogenesis. Although nicotinamide N-methyltransferase (NNMT) has been associated with obesity, how NNMT is regulated during adipogenesis, and the underlying regulatory mechanisms, remain undefined. In present study we used genetic and pharmacological approaches to elucidate the molecular signals driving NNMT activation and its role during adipogenesis. Firstly, we demonstrated that during the early phase of adipocyte differentiation NNMT is transactivated by CCAAT/Enhancer Binding Protein beta (CEBPB) in response to glucocorticoid (GC) induction. We found that Nnmt knockout, using CRISPR/Cas9 approach, impaired terminal adipogenesis by influencing the timing of cellular commitment and cell cycle exit during mitotic clonal expansion, as demonstrated by cell cycle analysis and RNA sequencing experiments. Biochemical and computational methods showed that a novel small molecule, called CC-410, stably binds to and highly specifically inhibits NNMT. CC-410 was, therefore, used to modulate protein activity during pre-adipocyte differentiation stages, demonstrating that, in line with the genetic approach, chemical inhibition of NNMT at the early stages of adipogenesis impairs terminal differentiation by deregulating the GC network. These congruent results conclusively demonstrate that NNMT is a key component of the GC-CEBP axis during the early stages of adipogenesis and could be a potential therapeutic target for both early-onset obesity and glucocorticoid-induced obesity

Integrative methylome-transcriptome analysis unravels cancer cell vulnerabilities in infant MLL-rearranged B cell acute lymphoblastic leukemia

B cell acute lymphoblastic leukemia (B-ALL) is the most common childhood cancer. As predicted by its prenatal origin, infant B-ALL (iB-ALL) shows an exceptionally silent DNA mutational landscape, suggesting that alternative epigenetic mechanisms may substantially contribute to its leukemogenesis. Here, we have integrated genome-wide DNA methylome and transcriptome data from 69 patients with de novo MLL-rearranged leukemia (MLLr) and non-MLLr iB-ALL leukemia uniformly treated according to the Interfant-99/06 protocol. iB-ALL methylome signatures display a plethora of common and specific alterations associated with chromatin states related to enhancer and transcriptional control in normal hematopoietic cells. DNA methylation, gene expression, and gene coexpression network analyses segregated MLLr away from non-MLLr iB-ALL and identified a coordinated and enriched expression of the AP-1 complex members FOS and JUN and RUNX factors in MLLr iB-ALL, consistent with the significant enrichment of hypomethylated CpGs in these genes. Integrative methylome-transcriptome analysis identified consistent cancer cell vulnerabilities, revealed a robust iB-ALL–specific gene expression–correlating dmCpG signature, and confirmed an epigenetic control of AP-1 and RUNX members in reshaping the molecular network of MLLr iB-ALL. Finally, pharmacological inhibition or functional ablation of AP-1 dramatically impaired MLLr-leukemic growth in vitro and in vivo using MLLr-iB-ALL patient–derived xenografts, providing rationale for new therapeutic avenues in MLLr-iB-ALL