Neuronal methylome reveals CREB-associated neuro-axonal impairment in multiple sclerosis
Adzemovic, Milena Z.
KAUST DepartmentBiological and Environmental Sciences and Engineering (BESE) Division
Online Publication Date2019-05-30
Print Publication Date2019-12
Permanent link to this recordhttp://hdl.handle.net/10754/656314
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AbstractBackground: Due to limited access to brain tissue, the precise mechanisms underlying neuro-axonal dysfunction in neurological disorders such as multiple sclerosis (MS) are largely unknown. In that context, profiling DNA methylation, which is a stable and cell type-specific regulatory epigenetic mark of genome activity, offers a unique opportunity to characterize the molecular mechanisms underpinning brain pathology in situ. We examined DNA methylation patterns of neuronal nuclei isolated from post-mortem brain tissue to infer processes that occur in neurons of MS patients. Results: We isolated subcortical neuronal nuclei from post-mortem white matter tissue of MS patients and non-neurological controls using flow cytometry. We examined bulk DNA methylation changes (total n = 29) and further disentangled true DNA methylation (5mC) from neuron-specific DNA hydroxymethylation (5hmC) (n = 17), using Illumina Infinium 450K arrays. We performed neuronal sub-type deconvolution using glutamate and GABA methylation profiles to further reduce neuronal sample heterogeneity. In total, we identified 2811 and 1534 significant (genome-wide adjusted P value < 0.05) differentially methylated and hydroxymethylated positions between MS patients and controls. We found striking hypo-5mC and hyper-5hmC changes occurring mainly within gene bodies, which correlated with reduced transcriptional activity, assessed using published RNAseq data from bulk brain tissue of MS patients and controls. Pathway analyses of the two cohorts implicated dysregulation of genes involved in axonal guidance and synaptic plasticity, with meta-analysis confirming CREB signalling as the most highly enriched pathway underlying these processes. We functionally investigated DNA methylation changes of CREB signalling-related genes by immunohistofluoresence of phosphorylated CREB in neurons from brain sections of a subcohort of MS patients and controls (n = 15). Notably, DNA methylation changes associated with a reduction of CREB activity in white matter neurons of MS patients compared to controls. Conclusions: Our data demonstrate that investigating 5mC and 5hmC modifications separately allows the discovery of a substantial fraction of changes occurring in neurons, which can escape traditional bisulfite-based DNA methylation analysis. Collectively, our findings indicate that neurons of MS patients acquire sustained hypo-5mC and hyper-5hmC, which may impair CREB-mediated neuro-axonal integrity, in turn relating to clinical symptoms.
CitationKular, L., Needhamsen, M., Adzemovic, M. Z., Kramarova, T., Gomez-Cabrero, D., Ewing, E., … Jagodic, M. (2019). Neuronal methylome reveals CREB-associated neuro-axonal impairment in multiple sclerosis. Clinical Epigenetics, 11(1). doi:10.1186/s13148-019-0678-1
SponsorsWe are grateful to A. van Vollenhoven for flow cytometry processing (Center for Molecular Medicine and Karolinska Institutet core facility), D. Fauvin for assistance with TrueMethyl and T. Morris for support in the BS/oxBS data pipeline. We thank R. Covacu for helpful feedback on data interpretation. We acknowledge GenomeScan/ServiceXS (Leiden, The Netherlands) for processing Illumina 450K array BS and oxBS data (cohort 2) and BEA core facility (Karolinska Institutet) for processing Illumina 450K array BS data (pilot samples and cohort 1). We thank the Multiple Sclerosis and Parkinson’s Tissue Bank (Imperial College London) for provision of brain tissue samples. Computations were performed on resources provided by SNIC through Uppsala Multidisciplinary Center for Advanced Computational Science (UPPMAX).
Funding This work was supported by grants from the Swedish Research Council, the Swedish Association for Persons with Neurological Disabilities, the Swedish Brain Foundation, the Swedish MS Foundation, Åsa Vilhelmsson Foundation, Petrus and Augusta Hedlunds Foundation, the Stockholm County Council (ALF project) and Karolinska Institutet. L. Kular was supported by fellowship from the Margaretha af Ugglas Foundation. D. Gomez-Cabrero and J. Tegner were supported by EU FP7 306000 STATegra.
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Kular, L., Needhamsen, M., Adzemovic, M., Kramarova, T., Gomez-Cabrero, D., Ewing, E., Piket, E., Tegnér, J., Beck, S., Piehl, F., Brundin, L., & Jagodic, M. (2019). Neuronal methylome reveals CREB-associated neuro-axonal impairment in multiple sclerosis. Figshare. https://doi.org/10.6084/M9.FIGSHARE.C.4524122.V1. DOI: 10.6084/m9.figshare.c.4524122.v1 Handle: 10754/664737
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