Publications by year
In Press
Smith RG, Pishva E, Shireby G, Smith AR, Roubroeks JAY, Hannon E, Wheildon G, Mastroeni D, Gasparoni G, Riemenschneider M, et al (In Press). A meta-analysis of epigenome-wide association studies in Alzheimer’s disease highlights novel differentially methylated loci across cortex.
Abstract:
A meta-analysis of epigenome-wide association studies in Alzheimer’s disease highlights novel differentially methylated loci across cortex
ABSTRACTEpigenome-wide association studies of Alzheimer’s disease have highlighted neuropathology-associated DNA methylation differences, although existing studies have been limited in sample size and utilized different brain regions. Here, we combine data from six DNA methylomic studies of Alzheimer’s disease (N=1,453 unique individuals) to identify differential methylation associated with Braak stage in different brain regions and across cortex. We identify 236 CpGs in the prefrontal cortex, 95 CpGs in the temporal gyrus and ten CpGs in the entorhinal cortex at Bonferroni significance, with none in the cerebellum. Our cross-cortex meta-analysis (N=1,408 donors) identifies 220 CpGs associated with neuropathology, annotated to 121 genes, of which 84 genes have not been previously reported at this significance threshold. We have replicated our findings using two further DNA methylomic datasets consisting of a further > 600 unique donors. The meta-analysis summary statistics are available in our online data resource (www.epigenomicslab.com/ad-meta-analysis/).
Abstract.
Dabin L, Guntoro F, Campbell T, Bélicard T, Smith AR, Smith RG, Raybould R, Schott JM, Lunnon K, Sarkies P, et al (In Press). Altered DNA methylation profiles in blood from patients with sporadic Creutzfeldt-Jakob disease.
Abstract:
Altered DNA methylation profiles in blood from patients with sporadic Creutzfeldt-Jakob disease
AbstractPrion diseases are fatal and transmissible neurodegenerative disorders caused by the misfolding and aggregation of prion protein. Although recent studies have implicated epigenetic variation in common neurodegenerative disorders, no study has yet explored their role in human prion diseases. Here we profiled genome-wide blood DNA methylation in the most common human prion disease, sporadic Creutzfeldt-Jakob disease (sCJD). Our case-control study (n=219), when accounting for differences in cell type composition between individuals, identified 38 probes at genome-wide significance (p < 1.24×0-7). Nine of these sites were taken forward in a replication study, performed in an independent case-control (n=186) cohort using pyrosequencing. Sites in or close to FKBP5, AIM2 (2 probes), UHRF1, KCNAB2, PRNP, ANK1 successfully replicated. The blood-based DNA methylation signal was tissue- and disease-specific, in that the replicated probe signals were unchanged in case-control studies using sCJD frontal-cortex (n=84), blood samples from patients with Alzheimer’s disease, and from inherited and acquired prion diseases. Machine learning algorithms using blood DNA methylation array profiles accurately distinguished sCJD patients and controls. Finally, we identified sites whose methylation levels associated with prolonged survival in sCJD patients. Altogether, this study has identified a peripheral DNA methylation signature of sCJD with a variety of potential biomarker applications.
Abstract.
Shireby GL, Davies JP, Francis PT, Burrage J, Walker EM, Neilson GWA, Dahir A, Thomas AJ, Love S, Smith RG, et al (In Press). Recalibrating the Epigenetic Clock: Implications for Assessing Biological Age in the Human Cortex.
Abstract:
Recalibrating the Epigenetic Clock: Implications for Assessing Biological Age in the Human Cortex
AbstractHuman DNA-methylation data have been used to develop biomarkers of ageing - referred to ‘epigenetic clocks’ - that have been widely used to identify differences between chronological age and biological age in health and disease including neurodegeneration, dementia and other brain phenotypes. Existing DNA methylation clocks are highly accurate in blood but are less precise when used in older samples or on brain tissue. We aimed to develop a novel epigenetic clock that performs optimally in human cortex tissue and has the potential to identify phenotypes associated with biological ageing in the brain. We generated an extensive dataset of human cortex DNA methylation data spanning the life-course (n = 1,397, ages = 1 to 104 years). This dataset was split into ‘training’ and ‘testing’ samples (training: n = 1,047; testing: n = 350). DNA methylation age estimators were derived using a transformed version of chronological age on DNA methylation at specific sites using elastic net regression, a supervised machine learning method. The cortical clock was subsequently validated in a novel human cortex dataset (n = 1,221, ages = 41 to 104 years) and tested for specificity in a large whole blood dataset (n = 1,175, ages = 28 to 98 years). We identified a set of 347 DNA methylation sites that, in combination optimally predict age in the human cortex. The sum of DNA methylation levels at these sites weighted by their regression coefficients provide the cortical DNA methylation clock age estimate. The novel clock dramatically out-performed previously reported clocks in additional cortical datasets. Our findings suggest that previous associations between predicted DNA methylation age and neurodegenerative phenotypes might represent false positives resulting from clocks not robustly calibrated to the tissue being tested and for phenotypes that become manifest in older ages. The age distribution and tissue type of samples included in training datasets need to be considered when building and applying epigenetic clock algorithms to human epidemiological or disease cohorts.
Abstract.
2022
Harvey J, Smith AR, Weymouth LS, Smith RG, Hubbard L, Bresner K, Pishva E, Williams N, Lunnon K, Creese B, et al (2022). An epigenome wide association study of sub‐phenotypes in Parkinson’s disease. Alzheimer's & Dementia, 18(S4).
MacBean LF, Smith AR, Smith RG, Lunnon K (2022). An integrated systems‐level analysis of the molecular changes resulting from systemic infections in Alzheimer’s disease. Alzheimer's & Dementia, 18(S4).
MacBean LF, Smith AR, Smith RG, Boche D, Lunnon K (2022). An integrated systems‐level analysis of the molecular changes resulting from systemic inflammation and amyloid‐beta immunisation in Alzheimer’s disease. Alzheimer's & Dementia, 18(S3).
Wheildon G, Weymouth LS, Smith AR, Smith RG, Troakes C, Al‐Sarraj S, Lunnon K (2022). DNA methylation profiling across brain regions in Huntington’s disease. Alzheimer's & Dementia, 18(S4).
Shireby G, Dempster E, Policicchio S, Smith RG, Pishva E, Chioza B, Davies JP, Burrage J, Lunnon K, Seiler-Vellame D, et al (2022). DNA methylation signatures of Alzheimer’s disease neuropathology in the cortex are primarily driven by variation in non-neuronal cell-types.
Shireby G, Dempster EL, Policicchio S, Smith RG, Pishva E, Chioza B, Davies JP, Burrage J, Lunnon K, Seiler Vellame D, et al (2022). DNA methylation signatures of Alzheimer’s disease neuropathology in the cortex are primarily driven by variation in non-neuronal cell-types.
Nature Communications,
13(1).
Abstract:
DNA methylation signatures of Alzheimer’s disease neuropathology in the cortex are primarily driven by variation in non-neuronal cell-types
AbstractAlzheimer’s disease (AD) is a chronic neurodegenerative disease characterized by the progressive accumulation of amyloid-beta and neurofibrillary tangles of tau in the neocortex. We profiled DNA methylation in two regions of the cortex from 631 donors, performing an epigenome-wide association study of multiple measures of AD neuropathology. We meta-analyzed our results with those from previous studies of DNA methylation in AD cortex (total n = 2013 donors), identifying 334 cortical differentially methylated positions (DMPs) associated with AD pathology including methylomic variation at loci not previously implicated in dementia. We subsequently profiled DNA methylation in NeuN+ (neuronal-enriched), SOX10+ (oligodendrocyte-enriched) and NeuN–/SOX10– (microglia- and astrocyte-enriched) nuclei, finding that the majority of DMPs identified in ‘bulk’ cortex tissue reflect DNA methylation differences occurring in non-neuronal cells. Our study highlights the power of utilizing multiple measures of neuropathology to identify epigenetic signatures of AD and the importance of characterizing disease-associated variation in purified cell-types.
Abstract.
Ma Y, Yu L, Olah M, Smith R, Oatman SR, Allen M, Pishva E, Zhang B, Menon V, Ertekin-Taner N, et al (2022). Epigenomic features related to microglia are associated with attenuated effect of APOE epsilon 4 on Alzheimer's disease risk in humans.
ALZHEIMERS & DEMENTIA,
18(4), 688-699.
Author URL.
Devall M, Soanes DM, Smith AR, Dempster EL, Smith RG, Burrage J, Iatrou A, Hannon E, Troakes C, Moore K, et al (2022). Genome-wide characterization of mitochondrial DNA methylation in human brain.
Front Endocrinol (Lausanne),
13Abstract:
Genome-wide characterization of mitochondrial DNA methylation in human brain.
BACKGROUND: There is growing interest in the role of DNA methylation in regulating the transcription of mitochondrial genes, particularly in brain disorders characterized by mitochondrial dysfunction. Here, we present a novel approach to interrogate the mitochondrial DNA methylome at single base resolution using targeted bisulfite sequencing. We applied this method to investigate mitochondrial DNA methylation patterns in post-mortem superior temporal gyrus and cerebellum brain tissue from seven human donors. RESULTS: We show that mitochondrial DNA methylation patterns are relatively low but conserved, with peaks in DNA methylation at several sites, such as within the D-LOOP and the genes MT-ND2, MT-ATP6, MT-ND4, MT-ND5 and MT-ND6, predominantly in a non-CpG context. The elevated DNA methylation we observe in the D-LOOP we validate using pyrosequencing. We identify loci that show differential DNA methylation patterns associated with age, sex and brain region. Finally, we replicate previously reported differentially methylated regions between brain regions from a methylated DNA immunoprecipitation sequencing study. CONCLUSIONS: We have annotated patterns of DNA methylation at single base resolution across the mitochondrial genome in human brain samples. Looking to the future this approach could be utilized to investigate the role of mitochondrial epigenetic mechanisms in disorders that display mitochondrial dysfunction.
Abstract.
Author URL.
MacBean LF, Smith AR, Smith RG, Lunnon K (2022). Investigating the molecular consequences of co‐morbidites in Alzheimer’s disease. Alzheimer's & Dementia, 18(S4).
Wheildon G, Smith AR, Soanes D, Smith RG, Moore K, O'Neill P, Morgan K, Thomas AJ, Francis PT, Love S, et al (2022). Targeted bisulfite sequencing analysis of candidate genes associated with Alzheimer’s disease. Alzheimer's & Dementia, 18(S4).
2021
Smith RG, Pishva E, Shireby G, Smith AR, Roubroeks JAY, Hannon E, Wheildon G, Mastroeni D, Gasparoni G, Riemenschneider M, et al (2021). A meta-analysis of epigenome-wide association studies in Alzheimer's disease highlights novel differentially methylated loci across cortex.
Nat Commun,
12(1).
Abstract:
A meta-analysis of epigenome-wide association studies in Alzheimer's disease highlights novel differentially methylated loci across cortex.
Epigenome-wide association studies of Alzheimer's disease have highlighted neuropathology-associated DNA methylation differences, although existing studies have been limited in sample size and utilized different brain regions. Here, we combine data from six DNA methylomic studies of Alzheimer's disease (N = 1453 unique individuals) to identify differential methylation associated with Braak stage in different brain regions and across cortex. We identify 236 CpGs in the prefrontal cortex, 95 CpGs in the temporal gyrus and ten CpGs in the entorhinal cortex at Bonferroni significance, with none in the cerebellum. Our cross-cortex meta-analysis (N = 1408 donors) identifies 220 CpGs associated with neuropathology, annotated to 121 genes, of which 84 genes have not been previously reported at this significance threshold. We have replicated our findings using two further DNA methylomic datasets consisting of a further >600 unique donors. The meta-analysis summary statistics are available in our online data resource ( www.epigenomicslab.com/ad-meta-analysis/ ).
Abstract.
Author URL.
MacBean LF, Smith RG, Smith AR, Nicoll JA, Boche D, Lunnon K (2021). An integrated systems-level analysis of the molecular changes resulting from systemic inflammation in Alzheimer's disease.
Alzheimers Dement,
17 Suppl 3Abstract:
An integrated systems-level analysis of the molecular changes resulting from systemic inflammation in Alzheimer's disease.
BACKGROUND: in the current study we have performed an Epigenome-Wide Association Study (EWAS) comparing bulk brain tissue in individuals with AD and non-demented controls with or without a systemic infection, followed by targeted validation in isolated microglia. METHOD: DNA methylation is being quantified from 300 post-mortem brain samples, 75 AD cases with infection, 75 AD cases without an infection, 75 non-demented controls with infection and 75 non-demented controls without an infection. DNA methylation of isolated microglia is also being quantified and gene expression data collected from the same samples to allow integration of epigenetic and transcriptomic data. RESULT: Statistical analyses performed using our established EWAS pipeline will identify significant differentially methylated positions. This is particularly expected to be observable in regions across genes implicated in disease pathology. CONCLUSION: Through cross-examination of the four study groups we will identify novel patterns of methylation in those suffering from neuroinflammation not previously observed, also providing critical insight into the underlying molecular changes that arise in microglia as a result of neuroinflammation.
Abstract.
Author URL.
Wheildon G, Smith AR, Smith RG, Lunnon K (2021). Targeted bisulfite sequencing analysis of candidate genes associated with Alzheimer's disease.
Alzheimers Dement,
17 Suppl 3Abstract:
Targeted bisulfite sequencing analysis of candidate genes associated with Alzheimer's disease.
BACKGROUND: the histopathological changes in Alzheimer's disease (AD), including extensive deposits of amyloid β plaques and neurofibrillary tangles occur many years before the onset of clinical symptoms and epigenetic processes such as DNA methylation may contribute to this delay. Recent epigenome-wide association studies (EWAS) have identified a number of loci in specific genes that show robust and reproducible alterations in DNA methylation changes in AD brain samples. However, the technologies used for these studies only assess a limited number of methylation sites in each gene and further analysis of methylation changes across the entire gene are required to determine the exact extent and pattern of methylation changes in disease. In this study we have performed targeted bisulfite sequencing and RNA sequencing in the Brains for Dementia Research (BDR) tissue sample resource, which is a highly characterised cohort containing tissue with a high degree of standardised pathological, clinical and administrative data available to allow comparative studies. METHODS: Prefrontal cortex brain samples from 96 individuals were selected from the BDR cohort and grouped by Braak stage (Control 0-II; mild cognitive impairment III-IV; AD V-VI). DNA and RNA was simultaneously extracted before next generation RNA-seq was performed on all 96 samples to analyse the AD transcriptome and data extracted for 30 genomic regions of interest identified from previous EWAS. Concurrently for the DNA, Agilent Sure Select target baits captured the same 30 target genomic regions that were bisulfite sequenced for a subset of 60 samples, allowing analysis of differentially methylated positions (DMPs). RESULTS: the exact location of DMPs within the targeted genomic regions were identified and compared between each group. Similarly, differential expression of mRNA transcripts encoded by these genes were also identified and related to methylation levels. CONCLUSION: This study builds on previous work that identified differential methylation in several genomic regions that were associated with Braak stage. By identifying the exact positions that are subjected to differential methylation and the potential impact these changes have on gene expression this work provides further evidence that dysregulation of methylation is associated with pathological changes in AD prefrontal cortex.
Abstract.
Author URL.
Smith AR, Smith RG, Macdonald R, Marzi SJ, Burrage J, Troakes C, Al-Sarraj S, Mill J, Lunnon K (2021). The histone modification H3K4me3 is altered at the <i>ANK1</i> locus in Alzheimer's disease brain.
Future Science OA,
7(4).
Abstract:
The histone modification H3K4me3 is altered at the ANK1 locus in Alzheimer's disease brain
Several epigenome-wide association studies of DNA methylation have highlighted altered DNA methylation in the ANK1 gene in Alzheimer's disease (AD) brain samples. However, no study has specifically examined ANK1 histone modifications in the disease. We use chromatin immunoprecipitation-qPCR to quantify tri-methylation at histone 3 lysine 4 (H3K4me3) and 27 (H3K27me3) in the ANK1 gene in entorhinal cortex from donors with high (n = 59) or low (n = 29) Alzheimer's disease pathology. We demonstrate decreased levels of H3K4me3, a marker of active gene transcription, with no change in H3K27me3, a marker of inactive genes. H3K4me3 is negatively correlated with DNA methylation in specific regions of the ANK1 gene. Our study suggests that the ANK1 gene shows altered epigenetic marks indicative of reduced gene activation in Alzheimer's disease.
Abstract.
2020
Dabin LC, Guntoro F, Campbell T, Bélicard T, Smith AR, Smith RG, Raybould R, Schott JM, Lunnon K, Sarkies P, et al (2020). Altered DNA methylation profiles in blood from patients with sporadic Creutzfeldt–Jakob disease.
Acta Neuropathologica,
140(6), 863-879.
Abstract:
Altered DNA methylation profiles in blood from patients with sporadic Creutzfeldt–Jakob disease
AbstractPrion diseases are fatal and transmissible neurodegenerative disorders caused by the misfolding and aggregation of prion protein. Although recent studies have implicated epigenetic variation in common neurodegenerative disorders, no study has yet explored their role in human prion diseases. Here we profiled genome-wide blood DNA methylation in the most common human prion disease, sporadic Creutzfeldt–Jakob disease (sCJD). Our case–control study (n = 219), when accounting for differences in cell type composition between individuals, identified 38 probes at genome-wide significance (p < 1.24 × 10–7). Nine of these sites were taken forward in a replication study, performed in an independent case–control (n = 186) cohort using pyrosequencing. Sites in or close to FKBP5, AIM2 (2 probes), UHRF1, KCNAB2. successfully replicated. The blood-based DNA methylation signal was tissue- and disease-specific, in that the replicated probe signals were unchanged in case–control studies using sCJD frontal-cortex (n = 84), blood samples from patients with Alzheimer’s disease, and from inherited and acquired prion diseases. Machine learning algorithms using blood DNA methylation array profiles accurately distinguished sCJD patients and controls. Finally, we identified sites whose methylation levels associated with prolonged survival in sCJD patients. Altogether, this study has identified a peripheral DNA methylation signature of sCJD with a variety of potential biomarker applications.
Abstract.
Roubroeks JAY, Smith AR, Smith RG, Pishva E, Ibrahim Z, Sattlecker M, Hannon EJ, Kłoszewska I, Mecocci P, Soininen H, et al (2020). An epigenome-wide association study of Alzheimer's disease blood highlights robust DNA hypermethylation in the HOXB6 gene.
Neurobiol Aging,
95, 26-45.
Abstract:
An epigenome-wide association study of Alzheimer's disease blood highlights robust DNA hypermethylation in the HOXB6 gene.
A growing number of epigenome-wide association studies have demonstrated a role for DNA methylation in the brain in Alzheimer's disease. With the aim of exploring peripheral biomarker potential, we have examined DNA methylation patterns in whole blood collected from 284 individuals in the AddNeuroMed study, which included 89 nondemented controls, 86 patients with Alzheimer's disease, and 109 individuals with mild cognitive impairment, including 38 individuals who progressed to Alzheimer's disease within 1 year. We identified significant differentially methylated regions, including 12 adjacent hypermethylated probes in the HOXB6 gene in Alzheimer's disease, which we validated using pyrosequencing. Using weighted gene correlation network analysis, we identified comethylated modules of genes that were associated with key variables such as APOE genotype and diagnosis. In summary, this study represents the first large-scale epigenome-wide association study of Alzheimer's disease and mild cognitive impairment using blood. We highlight the differences in various loci and pathways in early disease, suggesting that these patterns relate to cognitive decline at an early stage.
Abstract.
Author URL.
Ma Y, Yu L, Olah M, Smith R, Oatman SR, Allen M, Pishva E, Zhang B, Menon V, Ertekin-Taner N, et al (2020). EPIGENOMIC FEATURES RELATED TO MICROGLIA ARE ASSOCIATED WITH ATTENUATED EFFECT OF APOE ε4 ON ALZHEIMER'S DISEASE RISK IN HUMANS.
Alzheimers Dement,
16(Suppl 2).
Abstract:
EPIGENOMIC FEATURES RELATED TO MICROGLIA ARE ASSOCIATED WITH ATTENUATED EFFECT OF APOE ε4 ON ALZHEIMER'S DISEASE RISK IN HUMANS.
Not all APOE ε4 carriers who survive to advanced age develop Alzheimer's disease (AD); factors attenuating the risk of ε4 on AD may exist. Guided by the top ε4-attenuating signals from methylome-wide association analyses (N=572, ε4+ and ε4-) of neurofibrillary tangles and neuritic plaques, we conducted a meta-analysis for pathological AD within the ε4+ subgroups (N=235) across four independent collections of brains. Cortical RNA-seq and microglial morphology measurements were used in functional analyses. Three out of the four significant CpG dinucleotides were captured by one principle component (PC1), which interacts with ε4 on AD, and is associated with expression of innate immune genes and activated microglia. In ε4 carriers, reduction in each unit of PC1 attenuated the odds of AD by 58% (OR=2.39, 95%CI=[1.64,3.46], P=7.08x10-6). An epigenomic factor associated with a reduced proportion of activated microglia (microglial epigenomic factor 1) appears to attenuate the risk of ε4 on AD.
Abstract.
Author URL.
Ma Y, Yu L, Olah M, Smith R, Oatman SR, Allen M, Pishva E, Zhang B, Menon V, Ertekin-Taner N, et al (2020). Epigenomic features related to microglia are associated with attenuated effect of APOE ε4 on alzheimer’s disease risk in humans.
Tam HT (2020). Investigating APOE DNA methylation in Alzheimer's disease and its relationship to APOE ε4 genotype.
Abstract:
Investigating APOE DNA methylation in Alzheimer's disease and its relationship to APOE ε4 genotype
Alzheimer’s disease is a neurodegenerative condition that is the leading cause of dementia in the elderly, which affected close to 50 million people worldwide in 2017. The disease is characterised by the presence of amyloid beta plaques and neurofibrillary tangles of hyperphosphorylated tau protein in the brain. There are no disease-modifying treatments, largely because the mechanisms underlying disease and pathology are not understood. It is estimated that common genetic variants explain ~33% of disease incidence, although the mechanism behind their action is not clear. Polymorphisms in the APOE gene have been widely linked with AD, and the APOE ε4 variant is the greatest genetic risk factor for sporadic Alzheimer’s disease.
We hypothesise that epigenetic differences, namely DNA methylation, in APOE may contribute to Alzheimer’s disease aetiology and that this may also be related to APOE genotype. In this thesis we have investigated DNA methylation in prefrontal cortex and temporal gyrus tissue form individuals with varying degrees of pathology or disease using data from the Illumina 450K methylation array. We identified pathology-associated hypomethylation of APOE in four probes that reside in a CpG island in the 3’ untranslated region. Using a large cohort of pre-natal and post-natal brain tissue samples from individuals with methylation and matched genotype data we showed that methylation of one site in this region seems to be driven by APOE ε4 genotype in pre-natal development, although this was not observed in post-natal samples. We did however identify another loci in this island that showed hypomethylation with age in post-natal samples. In the promoter at the 5’ end of the gene we observed three adjacent loci near the transcription start site and one loci in the gene body that were significantly hypomethylated with advancing age in Alzheimer’s disease samples. Overall APOE ε4 genotype had little effect on APOE DNA methylation in the context of Alzheimer’s disease. This project is the first large-scale study of APOE DNA methylation with respect to Alzheimer’s disease diagnosis and pathology, age and APOE genotype and highlights the need for further work in to the role of APOE DNA methylation in disease aetiology.
Abstract.
Shireby GL, Davies JP, Francis PT, Burrage J, Walker EM, Neilson GWA, Dahir A, Thomas AJ, Love S, Smith RG, et al (2020). Recalibrating the epigenetic clock: implications for assessing biological age in the human cortex.
Brain,
143(12), 3763-3775.
Abstract:
Recalibrating the epigenetic clock: implications for assessing biological age in the human cortex.
Human DNA methylation data have been used to develop biomarkers of ageing, referred to as 'epigenetic clocks', which have been widely used to identify differences between chronological age and biological age in health and disease including neurodegeneration, dementia and other brain phenotypes. Existing DNA methylation clocks have been shown to be highly accurate in blood but are less precise when used in older samples or in tissue types not included in training the model, including brain. We aimed to develop a novel epigenetic clock that performs optimally in human cortex tissue and has the potential to identify phenotypes associated with biological ageing in the brain. We generated an extensive dataset of human cortex DNA methylation data spanning the life course (n = 1397, ages = 1 to 108 years). This dataset was split into 'training' and 'testing' samples (training: n = 1047; testing: n = 350). DNA methylation age estimators were derived using a transformed version of chronological age on DNA methylation at specific sites using elastic net regression, a supervised machine learning method. The cortical clock was subsequently validated in a novel independent human cortex dataset (n = 1221, ages = 41 to 104 years) and tested for specificity in a large whole blood dataset (n = 1175, ages = 28 to 98 years). We identified a set of 347 DNA methylation sites that, in combination, optimally predict age in the human cortex. The sum of DNA methylation levels at these sites weighted by their regression coefficients provide the cortical DNA methylation clock age estimate. The novel clock dramatically outperformed previously reported clocks in additional cortical datasets. Our findings suggest that previous associations between predicted DNA methylation age and neurodegenerative phenotypes might represent false positives resulting from clocks not robustly calibrated to the tissue being tested and for phenotypes that become manifest in older ages. The age distribution and tissue type of samples included in training datasets need to be considered when building and applying epigenetic clock algorithms to human epidemiological or disease cohorts.
Abstract.
Author URL.
2019
Smith AR, Smith RG, Burrage J, Troakes C, Al-Sarraj S, Kalaria RN, Sloan C, Robinson AC, Mill J, Lunnon K, et al (2019). A cross-brain regions study of ANK1 DNA methylation in different neurodegenerative diseases.
Neurobiology of Aging,
74, 70-76.
Abstract:
A cross-brain regions study of ANK1 DNA methylation in different neurodegenerative diseases
Recent epigenome-wide association studies in Alzheimer's disease have highlighted consistent robust neuropathology-associated DNA hypermethylation of the ankyrin 1 (ANK1) gene in the cortex. The extent to which altered ANK1 DNA methylation is also associated with other neurodegenerative diseases is not currently known. In the present study, we used bisulfite pyrosequencing to quantify DNA methylation across 8 CpG sites within a 118 bp region of the ANK1 gene across multiple brain regions in Alzheimer's disease, Vascular dementia, Dementia with Lewy bodies, Huntington's disease, and Parkinson's disease. We demonstrate disease-associated ANK1 hypermethylation in the entorhinal cortex in Alzheimer's disease, Huntington's disease, and Parkinson's disease, whereas in donors with Vascular dementia and Dementia with Lewy bodies, we observed elevated ANK1 DNA methylation only in individuals with coexisting Alzheimer's disease pathology. We did not observe any disease-associated differential ANK1 DNA methylation in the striatum in Huntington's disease or the substantia nigra in Parkinson's disease. Our data suggest that ANK1 is characterized by region and disease-specific differential DNA methylation in multiple neurodegenerative diseases.
Abstract.
Lardenoije R, Roubroeks J, Pishva E, Leber M, Wagner H, Iatrou A, Smith A, Smith R, Eijssen L, Kleineidam L, et al (2019). Alzheimer's Disease DNA (Hydroxy)Methylome in the Brain and Blood: Evidence for OXT Methylation as a Preclinical Marker.
Author URL.
Lardenoije R, Roubroeks JAY, Pishva E, Leber M, Wagner H, Iatrou A, Smith AR, Smith RG, Eijssen LMT, Kleineidam L, et al (2019). Alzheimer's disease-associated (hydroxy)methylomic changes in the brain and blood.
Clin Epigenetics,
11(1).
Abstract:
Alzheimer's disease-associated (hydroxy)methylomic changes in the brain and blood.
BACKGROUND: Late-onset Alzheimer's disease (AD) is a complex multifactorial affliction, the pathogenesis of which is thought to involve gene-environment interactions that might be captured in the epigenome. The present study investigated epigenome-wide patterns of DNA methylation (5-methylcytosine, 5mC) and hydroxymethylation (5-hydroxymethylcytosine, 5hmC), as well as the abundance of unmodified cytosine (UC), in relation to AD. RESULTS: We identified epigenetic differences in AD patients (n = 45) as compared to age-matched controls (n = 35) in the middle temporal gyrus, pertaining to genomic regions close to or overlapping with genes such as OXT (- 3.76% 5mC, pŠidák = 1.07E-06), CHRNB1 (+ 1.46% 5hmC, pŠidák = 4.01E-04), RHBDF2 (- 3.45% UC, pŠidák = 4.85E-06), and C3 (- 1.20% UC, pŠidák = 1.57E-03). In parallel, in an independent cohort, we compared the blood methylome of converters to AD dementia (n = 54) and non-converters (n = 42), at a preclinical stage. DNA methylation in the same region of the OXT promoter as found in the brain was found to be associated with subsequent conversion to AD dementia in the blood of elderly, non-demented individuals (+ 3.43% 5mC, pŠidák = 7.14E-04). CONCLUSIONS: the implication of genome-wide significant differential methylation of OXT, encoding oxytocin, in two independent cohorts indicates it is a promising target for future studies on early biomarkers and novel therapeutic strategies in AD.
Abstract.
Author URL.
Wong CCY, Smith RG, Hannon E, Assary E, Parikshak N, Prabhakar S, Geschwind D, Mill J (2019). CONVERGENT METHYLOMIC SIGNATURES BETWEEN AUTISM ASSOCIATED WITH DUPLICATIONS OF CHROMOSOME 15Q AND IDIOPATHIC AUTISM.
Author URL.
Wong CCY, Smith RG, Hannon E, Ramaswami G, Parikshak NN, Assary E, Troakes C, Poschmann J, Schalkwyk LC, Sun W, et al (2019). Genome-wide DNA methylation profiling identifies convergent molecular signatures associated with idiopathic and syndromic autism in post-mortem human brain tissue.
Hum Mol Genet,
28(13), 2201-2211.
Abstract:
Genome-wide DNA methylation profiling identifies convergent molecular signatures associated with idiopathic and syndromic autism in post-mortem human brain tissue.
Autism spectrum disorder (ASD) encompasses a collection of complex neuropsychiatric disorders characterized by deficits in social functioning, communication and repetitive behaviour. Building on recent studies supporting a role for developmentally moderated regulatory genomic variation in the molecular aetiology of ASD, we quantified genome-wide patterns of DNA methylation in 223 post-mortem tissues samples isolated from three brain regions [prefrontal cortex, temporal cortex and cerebellum (CB)] dissected from 43 ASD patients and 38 non-psychiatric control donors. We identified widespread differences in DNA methylation associated with idiopathic ASD (iASD), with consistent signals in both cortical regions that were distinct to those observed in the CB. Individuals carrying a duplication on chromosome 15q (dup15q), representing a genetically defined subtype of ASD, were characterized by striking differences in DNA methylationacross a discrete domain spanning an imprinted gene cluster within the duplicated region. In addition to the dramatic cis-effects on DNA methylation observed in dup15q carriers, we identified convergent methylomic signatures associated with both iASD and dup15q, reflecting the findings from previous studies of gene expression and H3K27ac. Cortical co-methylation network analysis identified a number of co-methylated modules significantly associated with ASD that are enriched for genomic regions annotated to genes involved in the immune system, synaptic signalling and neuronal regulation. Our study represents the first systematic analysis of DNA methylation associated with ASD across multiple brain regions, providing novel evidence for convergent molecular signatures associated with both idiopathic and syndromic autism.
Abstract.
Author URL.
Smith RG, Pishva E, Shireby G, Smith AR, Hannon E, Sharp AJ, Mastroeni D, Schalkwyk LC, Haroutunian V, Coleman PD, et al (2019). O4‐10‐06: INTEGRATED GENETIC‐EPIGENETIC ANALYSES OF ALZHEIMER'S DISEASE. Alzheimer's & Dementia, 15, p1261-p1261.
Smith AR, Smith RG, Pishva E, Hannon E, Roubroeks JAY, Burrage J, Troakes C, Al-Sarraj S, Sloan C, Mill J, et al (2019). Parallel profiling of DNA methylation and hydroxymethylation highlights neuropathology-associated epigenetic variation in Alzheimer's disease.
Clin Epigenetics,
11(1).
Abstract:
Parallel profiling of DNA methylation and hydroxymethylation highlights neuropathology-associated epigenetic variation in Alzheimer's disease.
BACKGROUND: Alzheimer's disease is a progressive neurodegenerative disorder that is hypothesized to involve epigenetic dysfunction. Previous studies of DNA modifications in Alzheimer's disease have been unable to distinguish between DNA methylation and DNA hydroxymethylation. DNA hydroxymethylation has been shown to be enriched in the human brain, although its role in Alzheimer's disease has not yet been fully explored. Here, we utilize oxidative bisulfite conversion, in conjunction with the Illumina Infinium Human Methylation 450K microarray, to identify neuropathology-associated differential DNA methylation and DNA hydroxymethylation in the entorhinal cortex. RESULTS: We identified one experiment-wide significant differentially methylated position residing in the WNT5B gene. Next, we investigated pathology-associated regions consisting of multiple adjacent loci. We identified one significant differentially hydroxymethylated region consisting of four probes spanning 104 bases in the FBXL16 gene. We also identified two significant differentially methylated regions: one consisting of two probes in a 93 base-pair region in the ANK1 gene and the other consisting of six probes in a 99-base pair region in the ARID5B gene. We also highlighted three regions that show alterations in unmodified cytosine: two probes in a 39-base pair region of ALLC, two probes in a 69-base pair region in JAG2, and the same six probes in ARID5B that were differentially methylated. Finally, we replicated significant ANK1 disease-associated hypermethylation and hypohydroxymethylation patterns across eight CpG sites in an extended 118-base pair region in an independent cohort using oxidative-bisulfite pyrosequencing. CONCLUSIONS: Our study represents the first epigenome-wide association study of both DNA methylation and hydroxymethylation in Alzheimer's disease entorhinal cortex. We demonstrate that previous estimates of DNA hypermethylation in ANK1 in Alzheimer's disease were underestimates as it is confounded by hypohydroxymethylation.
Abstract.
Author URL.
2018
Smith RG, Hannon E, De Jager PL, Chibnik L, Lott SJ, Condliffe D, Smith AR, Haroutunian V, Troakes C, Al-Sarraj S, et al (2018). Elevated DNA methylation across a 48-kb region spanning the HOXA gene cluster is associated with Alzheimer's disease neuropathology.
Alzheimers Dement,
14(12), 1580-1588.
Abstract:
Elevated DNA methylation across a 48-kb region spanning the HOXA gene cluster is associated with Alzheimer's disease neuropathology.
INTRODUCTION: Alzheimer's disease is a neurodegenerative disorder that is hypothesized to involve epigenetic dysregulation of gene expression in the brain. METHODS: We performed an epigenome-wide association study to identify differential DNA methylation associated with neuropathology in prefrontal cortex and superior temporal gyrus samples from 147 individuals, replicating our findings in two independent data sets (N = 117 and 740). RESULTS: We identify elevated DNA methylation associated with neuropathology across a 48-kb region spanning 208 CpG sites within the HOXA gene cluster. A meta-analysis of the top-ranked probe within the HOXA3 gene (cg22962123) highlighted significant hypermethylation across all three cohorts (P = 3.11 × 10-18). DISCUSSION: We present robust evidence for elevated DNA methylation associated with Alzheimer's disease neuropathology spanning the HOXA gene cluster on chromosome 7. These data add to the growing evidence highlighting a role for epigenetic variation in Alzheimer's disease, implicating the HOX gene family as a target for future investigation.
Abstract.
Author URL.
Wong CCY, Smith RG, Hannon E, Ramaswami G, Parikshak NN, Assary E, Troakes C, Poschmann J, Schalkwyk LC, Sun W, et al (2018). Genome-wide DNA methylation profiling identifies convergent molecular signatures associated with idiopathic and syndromic forms of autism in post-mortem human brain tissue.
Cecil CAM, Walton E, Jaffee SR, O'Connor T, Maughan B, Relton CL, Smith RG, McArdle W, Gaunt TR, Ouellet-Morin I, et al (2018). Neonatal DNA methylation and early-onset conduct problems: a genome-wide, prospective study.
Dev Psychopathol,
30(2), 383-397.
Abstract:
Neonatal DNA methylation and early-onset conduct problems: a genome-wide, prospective study.
Early-onset conduct problems (CP) are a key predictor of adult criminality and poor mental health. While previous studies suggest that both genetic and environmental risks play an important role in the development of early-onset CP, little is known about potential biological processes underlying these associations. In this study, we examined prospective associations between DNA methylation (cord blood at birth) and trajectories of CP (4-13 years), using data drawn from the Avon Longitudinal Study of Parents and Children. Methylomic variation at seven loci across the genome (false discovery rate < 0.05) differentiated children who go on to develop early-onset (n = 174) versus low (n = 86) CP, including sites in the vicinity of the monoglyceride lipase (MGLL) gene (involved in endocannabinoid signaling and pain perception). Subthreshold associations in the vicinity of three candidate genes for CP (monoamine oxidase a [MAOA], brain-derived neurotrophic factor [BDNF], and FK506 binding protein 5 [FKBP5]) were also identified. Within the early-onset CP group, methylation levels of the identified sites did not distinguish children who will go on to persist versus desist in CP behavior over time. Overall, we found that several of the identified sites correlated with prenatal exposures, and none were linked to known genetic methylation quantitative trait loci. Findings contribute to a better understanding of epigenetic patterns associated with early-onset CP.
Abstract.
Author URL.
2017
Smith RG, Lunnon K (2017). DNA Modifications and Alzheimer's Disease.
Adv Exp Med Biol,
978, 303-319.
Abstract:
DNA Modifications and Alzheimer's Disease.
Alzheimer's disease (AD) is a complex neurodegenerative disease, affecting millions of people worldwide. While a number of studies have focused on identifying genetic variants that contribute to the development and progression of late-onset AD, the majority of these only have a relatively small effect size. There are also a number of other risk factors, for example, age, gender, and other comorbidities; however, how these influence disease risk is not known. Therefore, in recent years, research has begun to investigate epigenetic mechanisms for a potential role in disease etiology. In this chapter, we discuss the current state of play for research into DNA modifications in AD, the most well studied being 5-methylcytosine (5-mC). We describe the earlier studies of candidate genes and global measures of DNA modifications in human AD samples, in addition to studies in mouse models of AD. We focus on recent epigenome-wide association studies (EWAS) in human AD, using microarray technology, examining a number of key study design issues pertinent to such studies. Finally, we discuss how new technological advances could further progress the research field.
Abstract.
Author URL.
Smith R, Lunnon K (2017). DNA Modifications and Alzheimer’s Disease. In (Ed)
Neuroepigenomics in Aging and Disease, Springer.
Abstract:
DNA Modifications and Alzheimer’s Disease
Abstract.
Roubroeks JAY, Smith RG, van den Hove DLA, Lunnon K (2017). Epigenetics and DNA methylomic profiling in Alzheimer's disease and other neurodegenerative diseases.
J Neurochem,
143(2), 158-170.
Abstract:
Epigenetics and DNA methylomic profiling in Alzheimer's disease and other neurodegenerative diseases.
Recent studies have suggested a role for epigenetic mechanisms in the complex etiology of various neurodegenerative diseases. In this review, we discuss advances that have been made toward understanding the role of epigenetic processes in neurodegenerative disorders, with a particular focus on Alzheimer's disease, where the most extensive studies have been undertaken to date. We provide a brief overview of DNA modifications, followed by a summarization of studies of DNA modifications in Alzheimer's disease and other neurodegenerative diseases.
Abstract.
Author URL.
Janecka M, Marzi SJ, Parsons MJ, Liu L, Paya-Cano JL, Smith RG, Fernandes C, Schalkwyk LC (2017). Genetic polymorphisms and their association with brain and behavioural measures in heterogeneous stock mice. Scientific Reports, 7(1).
Devall M, Smith RG, Jeffries A, Hannon E, Davies MN, Schalkwyk L, Mill J, Weedon M, Lunnon K (2017). Regional differences in mitochondrial DNA methylation in human post-mortem brain tissue.
Clin Epigenetics,
9Abstract:
Regional differences in mitochondrial DNA methylation in human post-mortem brain tissue.
BACKGROUND: DNA methylation is an important epigenetic mechanism involved in gene regulation, with alterations in DNA methylation in the nuclear genome being linked to numerous complex diseases. Mitochondrial DNA methylation is a phenomenon that is receiving ever-increasing interest, particularly in diseases characterized by mitochondrial dysfunction; however, most studies have been limited to the investigation of specific target regions. Analyses spanning the entire mitochondrial genome have been limited, potentially due to the amount of input DNA required. Further, mitochondrial genetic studies have been previously confounded by nuclear-mitochondrial pseudogenes. Methylated DNA Immunoprecipitation Sequencing is a technique widely used to profile DNA methylation across the nuclear genome; however, reads mapped to mitochondrial DNA are often discarded. Here, we have developed an approach to control for nuclear-mitochondrial pseudogenes within Methylated DNA Immunoprecipitation Sequencing data. We highlight the utility of this approach in identifying differences in mitochondrial DNA methylation across regions of the human brain and pre-mortem blood. RESULTS: We were able to correlate mitochondrial DNA methylation patterns between the cortex, cerebellum and blood. We identified 74 nominally significant differentially methylated regions (p
Abstract.
Author URL.
2016
Cecil CAM, Walton E, Smith RG, Viding E, McCrory EJ, Relton CL, Suderman M, Pingault J-B, McArdle W, Gaunt TR, et al (2016). DNA methylation and substance-use risk: a prospective, genome-wide study spanning gestation to adolescence.
Transl Psychiatry,
6(12).
Abstract:
DNA methylation and substance-use risk: a prospective, genome-wide study spanning gestation to adolescence.
Epigenetic processes have been implicated in addiction; yet, it remains unclear whether these represent a risk factor and/or a consequence of substance use. Here, we believe we conducted the first genome-wide, longitudinal study to investigate whether DNA methylation patterns in early life prospectively associate with substance use in adolescence. The sample comprised of 244 youth (51% female) from the Avon Longitudinal Study of Parents and Children (ALSPAC), with repeated assessments of DNA methylation (Illumina 450k array; cord blood at birth, whole blood at age 7) and substance use (tobacco, alcohol and cannabis use; age 14-18). We found that, at birth, epigenetic variation across a tightly interconnected genetic network (n=65 loci; q
Abstract.
Author URL.
Smith AR, Mill J, Smith RG, Lunnon K (2016). Elucidating novel dysfunctional pathways in Alzheimer's disease by integrating loci identified in genetic and epigenetic studies.
Neuroepigenetics,
6, 32-50.
Abstract:
Elucidating novel dysfunctional pathways in Alzheimer's disease by integrating loci identified in genetic and epigenetic studies
Alzheimer's disease is a complex neurodegenerative disorder. A large number of genome-wide association studies have been performed, which have been supplemented more recently by the first epigenome-wide association studies, leading to the identification of a number of novel loci altered in disease. Twin studies have shown monozygotic twin discordance for Alzheimer's disease (Gatz et al. 2006), leading to the conclusion that a combination of genetic and epigenetic mechanisms is likely to be involved in disease etiology (Lunnon & Mill, 2013). This review focuses on identifying overlapping pathways between published genome-wide association studies and epigenome-wide association studies, highlighting dysfunctional synaptic, lipid metabolism, plasma membrane/cytoskeleton, mitochondrial, and immune cell activation pathways. Identifying common pathways altered in genetic and epigenetic studies will aid our understanding of disease mechanisms and identify potential novel targets for pharmacological intervention.
Abstract.
Cecil CAM, Smith RG, Walton E, Mill J, McCrory EJ, Viding E (2016). Epigenetic signatures of childhood abuse and neglect: Implications for psychiatric vulnerability.
J Psychiatr Res,
83, 184-194.
Abstract:
Epigenetic signatures of childhood abuse and neglect: Implications for psychiatric vulnerability.
Childhood maltreatment is a key risk factor for poor mental and physical health. Recently, variation in epigenetic processes, such as DNA methylation, has emerged as a potential pathway mediating this association; yet, the extent to which different forms of maltreatment may be characterized by unique vs shared epigenetic signatures is currently unknown. In this study, we quantified DNA methylation across the genome in buccal epithelial cell samples from a high-risk sample of inner-city youth (n = 124; age = 16-24; 53% female), 68% of whom reported experiencing at least one form of maltreatment while growing up. Our analyses aimed to identify methylomic variation associated with exposure to five major types of childhood maltreatment. We found that: (i) maltreatment types differ in the extent to which they associate with methylomic variation, with physical exposures showing the strongest associations; (ii) many of the identified loci are annotated to genes previously implicated in stress-related outcomes, including psychiatric and physical disorders (e.g. GABBR1, GRIN2D, CACNA2D4, PSEN2); and (iii) based on gene ontology analyses, maltreatment types not only show unique methylation patterns enriched for specific biological processes (e.g. physical abuse and cardiovascular function), but also share a 'common' epigenetic signature enriched for biological processes related to neural development and organismal growth. A stringent set of sensitivity analyses were also run to identify high-confidence associations. Together, findings lend novel insights into epigenetic signatures of childhood abuse and neglect, point to novel potential biomarkers for future investigation and support a molecular link between maltreatment and poor health outcomes. Nevertheless, it will be important in future to replicate findings, as the use of cross-sectional data and high rates of polyvictimization in our study make it difficult to fully disentangle the shared vs unique epigenetic signatures of maltreatment types. Furthermore, studies will be needed to test the role of potential moderators in the identified associations, including age of onset and chronicity of maltreatment exposure.
Abstract.
Author URL.
Lunnon K, Hannon E, G.Smith R, Dempster E, Wong C, Burrage J, Troakes C, Al-Sarraj S, Kepa A, Schalkwyk L, et al (2016). Erratum to: Variation in 5-hydroxymethylcytosine across human cortex and cerebellum [Genome Biol. 2016, 17, 27]. Genome Biology, 17(1).
Smith AR, Smith RG, Condliffe D, Hannon E, Schalkwyk L, Mill J, Lunnon K (2016). Increased DNA methylation near TREM2 is consistently seen in the superior temporal gyrus in Alzheimer's disease brain.
Neurobiol Aging,
47, 35-40.
Abstract:
Increased DNA methylation near TREM2 is consistently seen in the superior temporal gyrus in Alzheimer's disease brain.
Although mutations within the TREM2 gene have been robustly associated with Alzheimer's disease, it is not known whether alterations in the regulation of this gene are also involved in pathogenesis. Here, we present data demonstrating increased DNA methylation in the superior temporal gyrus in Alzheimer's disease brain at a CpG site located 289 bp upstream of the transcription start site of the TREM2 gene in 3 independent study cohorts using 2 different technologies (Illumina Infinium 450K methylation beadchip and pyrosequencing). A meta-analysis across all 3 cohorts reveals consistent AD-associated hypermethylation (p = 3.47E-08). This study highlights that extending genetic studies of TREM2 in AD to investigate epigenetic changes may nominate additional mechanisms by which disruption to this gene increases risk.
Abstract.
Author URL.
Smith A, Smith R, Roubroeks J, Hannon E, Burrage J, Troakes C, van den Hove D, Mill J, Lunnon K (2016). O2‐06‐05: Hydroxymethylomic Profiling Implicates Cortical Deregulation of ANK1 and APP in the Alzheimer's Disease Brain. Alzheimer's & Dementia, 12(7S_Part_5), p240-p241.
Marzi SJ, Meaburn EL, Dempster EL, Lunnon K, Paya-Cano JL, Smith RG, Volta M, Troakes C, Schalkwyk LC, Mill J, et al (2016). Tissue-specific patterns of allelically-skewed DNA methylation.
Epigenetics,
11(1), 24-35.
Abstract:
Tissue-specific patterns of allelically-skewed DNA methylation.
While DNA methylation is usually thought to be symmetrical across both alleles, there are some notable exceptions. Genomic imprinting and X chromosome inactivation are two well-studied sources of allele-specific methylation (ASM), but recent research has indicated a more complex pattern in which genotypic variation can be associated with allelically-skewed DNA methylation in cis. Given the known heterogeneity of DNA methylation across tissues and cell types we explored inter- and intra-individual variation in ASM across several regions of the human brain and whole blood from multiple individuals. Consistent with previous studies, we find widespread ASM with > 4% of the ∼220,000 loci interrogated showing evidence of allelically-skewed DNA methylation. We identify ASM flanking known imprinted regions, and show that ASM sites are enriched in DNase I hypersensitivity sites and often located in an extended genomic context of intermediate DNA methylation. We also detect examples of genotype-driven ASM, some of which are tissue-specific. These findings contribute to our understanding of the nature of differential DNA methylation across tissues and have important implications for genetic studies of complex disease. As a resource to the community, ASM patterns across each of the tissues studied are available in a searchable online database: http://epigenetics.essex.ac.uk/ASMBrainBlood.
Abstract.
Author URL.
Lunnon K, Hannon E, Smith RG, Dempster E, Wong C, Burrage J, Troakes C, Al-Sarraj S, Kepa A, Schalkwyk L, et al (2016). Variation in 5-hydroxymethylcytosine across human cortex and cerebellum.
Genome Biol,
17Abstract:
Variation in 5-hydroxymethylcytosine across human cortex and cerebellum.
BACKGROUND: the most widely utilized approaches for quantifying DNA methylation involve the treatment of genomic DNA with sodium bisulfite; however, this method cannot distinguish between 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC). Previous studies have shown that 5hmC is enriched in the brain, although little is known about its genomic distribution and how it differs between anatomical regions and individuals. In this study, we combine oxidative bisulfite (oxBS) treatment with the Illumina Infinium 450K BeadArray to quantify genome-wide patterns of 5hmC in two distinct anatomical regions of the brain from multiple individuals. RESULTS: We identify 37,145 and 65,563 sites passing our threshold for detectable 5hmC in the prefrontal cortex and cerebellum respectively, with 23,445 loci common across both brain regions. Distinct patterns of 5hmC are identified in each brain region, with notable differences in the genomic location of the most hydroxymethylated loci between these brain regions. Tissue-specific patterns of 5hmC are subsequently confirmed in an independent set of prefrontal cortex and cerebellum samples. CONCLUSIONS: This study represents the first systematic analysis of 5hmC in the human brain, identifying tissue-specific hydroxymethylated positions and genomic regions characterized by inter-individual variation in DNA hydroxymethylation. This study demonstrates the utility of combining oxBS-treatment with the Illumina 450k methylation array to systematically quantify 5hmC across the genome and the potential utility of this approach for epigenomic studies of brain disorders.
Abstract.
Author URL.
2015
Lunnon K, Smith RG, Cooper I, Greenbaum L, Mill J, Beeri MS (2015). Blood methylomic signatures of presymptomatic dementia in elderly subjects with type 2 diabetes mellitus.
Neurobiology of Aging,
36(3), 1600.e1-1600.e4.
Abstract:
Blood methylomic signatures of presymptomatic dementia in elderly subjects with type 2 diabetes mellitus
Due to an aging population, the incidence of dementia is steadily rising. The ability to identify early markers in blood, which appear before the onset of clinical symptoms is of considerable interest to allow early intervention, particularly in "high risk" groups such as those with type 2 diabetes. Here, we present a longitudinal study of genome-wide DNA methylation in whole blood from 18 elderly individuals with type 2 diabetes who developed presymptomatic dementia within an 18-month period following baseline assessment and 18 age-, sex-, and education-matched controls who maintained normal cognitive function. We identified a significant overlap in methylomic differences between groups at baseline and follow-up, with 8 CpG sites being consistently differentially methylated above our nominal significance threshold before symptoms at baseline and at 18months follow up, after a diagnosis of presymptomatic dementia. Finally, we report a significant overlap between DNA methylation differences identified in converters, only after they develop symptoms of dementia, with differences at the same loci in blood samples from patients with clinically diagnosed Alzheimer's disease compared with unaffected control subjects.
Abstract.
Lunnon K, Smith RG, Cooper I, Greenbaum L, Mill J, Beeri MS (2015). Blood methylomic signatures of presymptomatic dementia in elderly subjects with type 2 diabetes mellitus.
Neurobiol Aging,
36(3), 1600.e1-1600.e4.
Abstract:
Blood methylomic signatures of presymptomatic dementia in elderly subjects with type 2 diabetes mellitus.
Due to an aging population, the incidence of dementia is steadily rising. The ability to identify early markers in blood, which appear before the onset of clinical symptoms is of considerable interest to allow early intervention, particularly in "high risk" groups such as those with type 2 diabetes. Here, we present a longitudinal study of genome-wide DNA methylation in whole blood from 18 elderly individuals with type 2 diabetes who developed presymptomatic dementia within an 18-month period following baseline assessment and 18 age-, sex-, and education-matched controls who maintained normal cognitive function. We identified a significant overlap in methylomic differences between groups at baseline and follow-up, with 8 CpG sites being consistently differentially methylated above our nominal significance threshold before symptoms at baseline and at 18 months follow up, after a diagnosis of presymptomatic dementia. Finally, we report a significant overlap between DNA methylation differences identified in converters, only after they develop symptoms of dementia, with differences at the same loci in blood samples from patients with clinically diagnosed Alzheimer's disease compared with unaffected control subjects.
Abstract.
Author URL.
Spiers H, Bray NJ, Hannon E, Schalkwyk LC, Wong CC, Pidsley R, Smith RG, Mill J (2015). Dynamic and sex-specific changes in DNA methylation during human fetal brain development.
Int J Dev Neurosci,
47(Pt A), 50-51.
Author URL.
Janecka M, Manduca A, Servadio M, Trezza V, Smith R, Mill J, Schalkwyk LC, Reichenberg A, Fernandes C (2015). Effects of advanced paternal age on trajectories of social behavior in offspring.
Genes Brain Behav,
14(6), 443-453.
Abstract:
Effects of advanced paternal age on trajectories of social behavior in offspring.
Our study is the first investigation of the effects of advanced paternal age (APA) on the developmental trajectory of social behavior in rodent offspring. Given the strong epidemiological association between APA and sexually dimorphic neurodevelopmental disorders that are characterized by abnormalities in social behavior (autism, schizophrenia), we assessed sociability in male and female inbred mice (C57BL/6J) across postnatal development (N = 104) in relation to paternal age. We found differences in early social behavior in both male and female offspring of older breeders, with differences in this social domain persisting into adulthood in males only. We showed that these social deficits were not present in the fathers of these offspring, confirming a de novo origin of an altered social trajectory in the offspring generation. Our results, highly novel in rodent research, support the epidemiological observations in humans and provide evidence for a causal link between APA, age-related changes in the paternal sperm DNA and neurodevelopmental disorders in their offspring.
Abstract.
Author URL.
Spiers H, Hannon E, Schalkwyk LC, Smith R, Wong CCY, O'Donovan MC, Bray NJ, Mill J (2015). Methylomic trajectories across human fetal brain development.
Genome Res,
25(3), 338-352.
Abstract:
Methylomic trajectories across human fetal brain development.
Epigenetic processes play a key role in orchestrating transcriptional regulation during development. The importance of DNA methylation in fetal brain development is highlighted by the dynamic expression of de novo DNA methyltransferases during the perinatal period and neurodevelopmental deficits associated with mutations in the methyl-CpG binding protein 2 (MECP2) gene. However, our knowledge about the temporal changes to the epigenome during fetal brain development has, to date, been limited. We quantified genome-wide patterns of DNA methylation at ∼ 400,000 sites in 179 human fetal brain samples (100 male, 79 female) spanning 23 to 184 d post-conception. We identified highly significant changes in DNA methylation across fetal brain development at >7% of sites, with an enrichment of loci becoming hypomethylated with fetal age. Sites associated with developmental changes in DNA methylation during fetal brain development were significantly underrepresented in promoter regulatory regions but significantly overrepresented in regions flanking CpG islands (shores and shelves) and gene bodies. Highly significant differences in DNA methylation were observed between males and females at a number of autosomal sites, with a small number of regions showing sex-specific DNA methylation trajectories across brain development. Weighted gene comethylation network analysis (WGCNA) revealed discrete modules of comethylated loci associated with fetal age that are significantly enriched for genes involved in neurodevelopmental processes. This is, to our knowledge, the most extensive study of DNA methylation across human fetal brain development to date, confirming the prenatal period as a time of considerable epigenomic plasticity.
Abstract.
Author URL.
Lunnon K, Smith R, Hannon E, Smith A, Schalkwyk LC, Haroutunian V, Mill J (2015). O3‐05‐01: Systems‐level evidence for epigenetic dysfunction in Alzheimer's disease.
2014
Cecil CAM, Lysenko LJ, Jaffee SR, Pingault J-B, Smith RG, Relton CL, Woodward G, McArdle W, Mill J, Barker ED, et al (2014). Environmental risk, Oxytocin Receptor Gene (OXTR) methylation and youth callous-unemotional traits: a 13-year longitudinal study.
Mol Psychiatry,
19(10), 1071-1077.
Abstract:
Environmental risk, Oxytocin Receptor Gene (OXTR) methylation and youth callous-unemotional traits: a 13-year longitudinal study.
Youth with high callous-unemotional traits (CU) are at risk for early-onset and persistent conduct problems. Research suggests that there may be different developmental pathways to CU (genetic/constitutional vs environmental), and that the absence or presence of co-occurring internalizing problems is a key marker. However, it is unclear whether such a distinction is valid. Intermediate phenotypes such as DNA methylation, an epigenetic modification regulating gene expression, may help to clarify etiological pathways. This is the first study to examine prospective inter-relationships between environmental risk (prenatal/postnatal) and DNA methylation (birth, age 7 and 9) in the prediction of CU (age 13), for youth low vs high in internalizing problems. We focused on DNA methylation in the vicinity of the oxytocin receptor (OXTR) gene as it has been previously implicated in CU. Participants were 84 youth with early-onset and persistent conduct problems drawn from the Avon Longitudinal Study of Parents and Children. For youth with low internalizing problems (46%), we found that (i) OXTR methylation at birth associated with higher CU (age 13) as well as decreased experience of victimization during childhood (evocative epigenetic-environment correlation; birth-age 7), (ii) higher prenatal parental risks (maternal psychopathology, criminal behaviors, substance use) associated with higher OXTR methylation at birth and (iii) OXTR methylation levels were more stable across time (birth-age 9). In contrast, for youth with high internalizing problems, CU were associated with prenatal risks of an interpersonal nature (that is, intimate partner violence, family conflict) but not OXTR methylation. Findings support the existence of distinct developmental pathways to CU.
Abstract.
Author URL.
Lunnon K, Smith R, Hannon E, De Jager PL, Srivastava G, Volta M, Troakes C, Al-Sarraj S, Burrage J, Macdonald R, et al (2014). Methylomic profiling implicates cortical deregulation of ANK1 in Alzheimer's disease.
Nat Neurosci,
17(9), 1164-1170.
Abstract:
Methylomic profiling implicates cortical deregulation of ANK1 in Alzheimer's disease.
Alzheimer's disease (AD) is a chronic neurodegenerative disorder that is characterized by progressive neuropathology and cognitive decline. We performed a cross-tissue analysis of methylomic variation in AD using samples from four independent human post-mortem brain cohorts. We identified a differentially methylated region in the ankyrin 1 (ANK1) gene that was associated with neuropathology in the entorhinal cortex, a primary site of AD manifestation. This region was confirmed as being substantially hypermethylated in two other cortical regions (superior temporal gyrus and prefrontal cortex), but not in the cerebellum, a region largely protected from neurodegeneration in AD, or whole blood obtained pre-mortem from the same individuals. Neuropathology-associated ANK1 hypermethylation was subsequently confirmed in cortical samples from three independent brain cohorts. This study represents, to the best of our knowledge, the first epigenome-wide association study of AD employing a sequential replication design across multiple tissues and highlights the power of this approach for identifying methylomic variation associated with complex disease.
Abstract.
Author URL.
Lunnon K, Smith R, Hannon E, Volta M, Troakes C, Haroutunian V, Katsel P, Al‐Sarraj S, Lovestone S, Schalkwyk L, et al (2014). O3‐04‐03: CROSS‐TISSUE METHYLOMIC PROFILING IN ALZHEIMER'S DISEASE.
Lunnon K, Smith RG, Cooper I, Greenbaum L, Mill J, Beeri MS (2014). TEMPORARY REMOVAL: Blood methylomic signatures of presymptomatic dementia in elderly subjects with type 2 diabetes mellitus.
Neurobiol Aging Author URL.
Smith RG, Fernandes C, Kember R, Schalkwyk LC, Buxbaum J, Reichenberg A, Mill J (2014). Transcriptomic changes in the frontal cortex associated with paternal age.
Mol Autism,
5(1).
Abstract:
Transcriptomic changes in the frontal cortex associated with paternal age.
BACKGROUND: Advanced paternal age is robustly associated with several human neuropsychiatric disorders, particularly autism. The precise mechanism(s) mediating the paternal age effect are not known, but they are thought to involve the accumulation of de novo (epi)genomic alterations. In this study we investigate differences in the frontal cortex transcriptome in a mouse model of advanced paternal age. FINDINGS: Transcriptomic profiling was undertaken for medial prefrontal cortex tissue dissected from the male offspring of young fathers (2 month old, 4 sires, n = 16 offspring) and old fathers (10 month old, 6 sires, n = 16 offspring) in a mouse model of advancing paternal age. We found a number of differentially expressed genes in the offspring of older fathers, many previously implicated in the aetiology of autism. Pathway analysis highlighted significant enrichment for changes in functional networks involved in inflammation and inflammatory disease, which are also implicated in autism. CONCLUSIONS: We observed widespread alterations to the transcriptome associated with advanced paternal age with an enrichment of genes associated with inflammation, an interesting observation given previous evidence linking the immune system to several neuropsychiatric disorders including autism.
Abstract.
Author URL.
2013
Smith RG, Reichenberg A, Kember RL, Buxbaum JD, Schalkwyk LC, Fernandes C, Mill J (2013). Advanced paternal age is associated with altered DNA methylation at brain-expressed imprinted loci in inbred mice: implications for neuropsychiatric disease.
Mol Psychiatry,
18(6), 635-636.
Author URL.
Powell TR, Smith RG, Hackinger S, Schalkwyk LC, Uher R, McGuffin P, Mill J, Tansey KE (2013). DNA methylation in interleukin-11 predicts clinical response to antidepressants in GENDEP.
Transl Psychiatry,
3(9).
Abstract:
DNA methylation in interleukin-11 predicts clinical response to antidepressants in GENDEP.
Transcriptional differences in interleukin-11 (IL11) after antidepressant treatment have been found to correspond to clinical response in major depressive disorder (MDD) patients. Expression differences were partly mediated by a single-nucleotide polymorphism (rs1126757), identified as a predictor of antidepressant response as part of a genome-wide association study. Here we attempt to identify whether DNA methylation, another baseline factor known to affect transcription factor binding, might also predict antidepressant response, using samples collected from the Genome-based Therapeutic Drugs for Depression project (GENDEP). DNA samples from 113 MDD individuals from the GENDEP project, who were treated with either escitalopram (n=80) or nortriptyline (n=33) for 12 weeks, were randomly selected. Percentage change in Montgomery-Åsberg Depression Rating Scale scores between baseline and week 12 were utilized as our measure of antidepressant response. The Sequenom EpiTYPER platform was used to assess DNA methylation across the only CpG island located in the IL11 gene. Regression analyses were then used to explore the relationship between CpG unit methylation and antidepressant response. We identified a CpG unit predictor of general antidepressant response, a drug by CpG unit interaction predictor of response, and a CpG unit by rs1126757 interaction predictor of antidepressant response. The current study is the first to investigate the potential utility of pharmaco-epigenetic biomarkers for the prediction of antidepressant response. Our results suggest that DNA methylation in IL11 might be useful in identifying those patients likely to respond to antidepressants, and if so, the best drug suited to each individual.
Abstract.
Author URL.
Hodgson K, Tansey K, Dernovšek MZ, Hauser J, Henigsberg N, Maier W, Mors O, Placentino A, Rietschel M, Souery D, et al (2013). Genetic differences in cytochrome P450 enzymes and antidepressant treatment response. Journal of Psychopharmacology, 28(2), 133-141.
2012
Pidsley R, Fernandes C, Viana J, Paya-Cano JL, Liu L, Smith RG, Schalkwyk LC, Mill J (2012). DNA methylation at the Igf2/H19 imprinting control region is associated with cerebellum mass in outbred mice.
Mol Brain,
5Abstract:
DNA methylation at the Igf2/H19 imprinting control region is associated with cerebellum mass in outbred mice.
BACKGROUND: Insulin-like growth factor 2 (Igf2) is a paternally expressed imprinted gene regulating fetal growth, playing an integral role in the development of many tissues including the brain. The parent-of-origin specific expression of Igf2 is largely controlled by allele-specific DNA methylation at CTCF-binding sites in the imprinting control region (ICR), located immediately upstream of the neighboring H19 gene. Previously we reported evidence of a negative correlation between DNA methylation in this region and cerebellum weight in humans. RESULTS: We quantified cerebellar DNA methylation across all four CTCF binding sites spanning the murine Igf2/H19 ICR in an outbred population of Heterogeneous Stock (HS) mice (n = 48). We observe that DNA methylation at the second and third CTCF binding sites in the Igf2/H19 ICR shows a negative relationship with cerebellar mass, reflecting the association observed in human post-mortem cerebellum tissue. CONCLUSIONS: Given the important role of the cerebellum in motor control and cognition, and the link between structural cerebellar abnormalities and neuropsychiatric phenotypes, the identification of epigenetic factors associated with cerebellum growth and development may provide important insights about the etiology of psychiatric disorders.
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2011
Huezo-Diaz P, Perroud N, Spencer EP, Smith R, Sim S, Virding S, Uher R, Gunasinghe C, Gray J, Campbell D, et al (2011). CYP2C19 genotype predicts steady state escitalopram concentration in GENDEP. Journal of Psychopharmacology, 26(3), 398-407.
Smith R, Mill J (2011). Epigenetics and Chronic Diseases: an Overview. In (Ed) Epigenetic Aspects of Chronic Diseases, Springer London, 1-20.
2010
Schalkwyk LC, Meaburn EL, Smith R, Dempster EL, Jeffries AR, Davies MN, Plomin R, Mill J (2010). Allelic skewing of DNA methylation is widespread across the genome.
Am J Hum Genet,
86(2), 196-212.
Abstract:
Allelic skewing of DNA methylation is widespread across the genome.
DNA methylation is assumed to be complementary on both alleles across the genome, although there are exceptions, notably in regions subject to genomic imprinting. We present a genome-wide survey of the degree of allelic skewing of DNA methylation with the aim of identifying previously unreported differentially methylated regions (DMRs) associated primarily with genomic imprinting or DNA sequence variation acting in cis. We used SNP microarrays to quantitatively assess allele-specific DNA methylation (ASM) in amplicons covering 7.6% of the human genome following cleavage with a cocktail of methylation-sensitive restriction enzymes (MSREs). Selected findings were verified using bisulfite-mapping and gene-expression analyses, subsequently tested in a second tissue from the same individuals, and replicated in DNA obtained from 30 parent-child trios. Our approach detected clear examples of ASM in the vicinity of known imprinted loci, highlighting the validity of the method. In total, 2,704 (1.5%) of our 183,605 informative and stringently filtered SNPs demonstrate an average relative allele score (RAS) change > or =0.10 following MSRE digestion. In agreement with previous reports, the majority of ASM ( approximately 90%) appears to be cis in nature, and several examples of tissue-specific ASM were identified. Our data show that ASM is a widespread phenomenon, with >35,000 such sites potentially occurring across the genome, and that a spectrum of ASM is likely, with heterogeneity between individuals and across tissues. These findings impact our understanding about the origin of individual phenotypic differences and have implications for genetic studies of complex disease.
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Keers R, Uher R, Huezo-Diaz P, Smith R, Jaffee S, Rietschel M, Henigsberg N, Kozel D, Mors O, Maier W, et al (2010). Interaction between serotonin transporter gene variants and life events predicts response to antidepressants in the GENDEP project. The Pharmacogenomics Journal, 11(2), 138-145.
Drago A, Serretti A, Smith R, Huezo-Diaz P, Malitas P, Albani D, De Ronchi D, Pae C-U, Aitchison KJ (2010). No association between genetic markers in BDNF gene and lithium prophylaxis in a Greek sample. International Journal of Psychiatry in Clinical Practice, 14(2), 154-157.
2009
Smith RG, Kember RL, Mill J, Fernandes C, Schalkwyk LC, Buxbaum JD, Reichenberg A (2009). Advancing paternal age is associated with deficits in social and exploratory behaviors in the offspring: a mouse model.
PLoS One,
4(12).
Abstract:
Advancing paternal age is associated with deficits in social and exploratory behaviors in the offspring: a mouse model.
BACKGROUND: Accumulating evidence from epidemiological research has demonstrated an association between advanced paternal age and risk for several psychiatric disorders including autism, schizophrenia and early-onset bipolar disorder. In order to establish causality, this study used an animal model to investigate the effects of advanced paternal age on behavioural deficits in the offspring. METHODS: C57BL/6J offspring (n = 12 per group) were bred from fathers of two different ages, 2 months (young) and 10 months (old), and mothers aged 2 months (n = 6 breeding pairs per group). Social and exploratory behaviors were examined in the offspring. PRINCIPAL FINDINGS: the offspring of older fathers were found to engage in significantly less social (p = 0.02) and exploratory (p = 0.02) behaviors than the offspring of younger fathers. There were no significant differences in measures of motor activity. CONCLUSIONS: Given the well-controlled nature of this study, this provides the strongest evidence for deleterious effects of advancing paternal age on social and exploratory behavior. De-novo chromosomal changes and/or inherited epigenetic changes are the most plausible explanatory factors.
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Author URL.
Perroud N, Aitchison KJ, Uher R, Smith R, Huezo-Diaz P, Marusic A, Maier W, Mors O, Placentino A, Henigsberg N, et al (2009). Genetic Predictors of Increase in Suicidal Ideation During Antidepressant Treatment in the GENDEP Project. Neuropsychopharmacology, 34(12), 2517-2528.
Uher R, Huezo-Diaz P, Perroud N, Smith R, Rietschel M, Mors O, Hauser J, Maier W, Kozel D, Henigsberg N, et al (2009). Genetic predictors of response to antidepressants in the GENDEP project. The Pharmacogenomics Journal, 9(4), 225-233.
Huezo-Diaz P, Uher R, Smith R, Rietschel M, Henigsberg N, Marušiˇ A, Mors O, Maier W, Hauser J, Souery D, et al (2009). Moderation of antidepressant response by the serotonin transporter gene. The British Journal of Psychiatry, 195(1), 30-38.
