Parkinson’s disease (PD) pathogenesis may involve the epigenetic control of enhancers that modify neuronal functions. Here, we comprehensively examine DNA methylation at enhancers,

genome-wide, in neurons of patients with PD and of control individuals. We find a widespread increase in cytosine modifications at enhancers in PD neurons, which is partly explained by

elevated hydroxymethylation levels. In particular, patients with PD exhibit an epigenetic and transcriptional upregulation of TET2, a master-regulator of cytosine modification status. TET2

depletion in a neuronal cell model results in cytosine modification changes that are reciprocal to those observed in PD neurons. Moreover, Tet2 inactivation in mice fully prevents nigral

All sequencing data generated in this study are available from the NCBI Gene Expression Omnibus (GEO) database under the accession number GSE136010. DNA methylation padlock-seq and RNA-seq

data used in this study are publicly available under the GEO accession number GSE135037. Hi-C data are available at GSM2322542. RNA-seq data from Parkinson’s disease models in Extended Data

Fig. 7a–d are available under the GEO accession numbers GSE54795, GSE108370, GSE125239 and GSE130752.

https://github.com/LeeLMarshall/Epigenomic-analysis-of-Parkinson-s-disease-neurons-identifies-Tet2-loss-as-neuroprotectiveSource data are provided with this paper.

Code used for the analyses in this study is provided in the Supplementary Software and is available at

https://github.com/LeeLMarshall/Epigenomic-analysis-of-Parkinson-s-disease-neurons-identifies-Tet2-loss-as-neuroprotective.

We thank the Van Andel Institute Flow Cytometry, Genomics, Bioinformatics and Biostatistics, Pathology and Vivarium Cores. We also thank the CAMH Sequencing Facility. We thank the

Parkinson’s UK Brain Bank, the NIH NeuroBioBank and the Michigan Brain Bank for the brain tissue provided. This work was funded by a Department of Defense grant (grant no. W81XWH1810512) and

a VAI Innovation Award to V.L. V.L. is also supported by grants from the National Institute of Neurological Disorders and Stroke (grant no. 1R21NS112614-01, grant no. 1R01NS114409-01A1 and

grant no. 1R01NS113894-01A1), the Farmer Family Foundation Parkinson’s Research Initiative and a Gibby & Friends vs. Parky Award. S.J. and M.W. are supported by the Richard and Helen DeVos

Foundation.

Center for Neurodegenerative Science, Van Andel Institute, Grand Rapids, MI, USA

Lee L. Marshall, Bryan A. Killinger, Elizabeth Ensink, Peipei Li, Katie X. Li, Noah Lubben, Xinhe Wang, Juozas Gordevicius, Gerhard A. Coetzee, Jiyan Ma & Viviane Labrie

Department of Neurological Sciences, Rush Medical Center, Chicago, IL, USA

Center for Epigenetics, Van Andel Institute, Grand Rapids, MI, USA

DeVos Cardiovascular Research Program Van Andel Institute-Spectrum Health, Grand Rapids, MI, USA

Institute of Biotechnology, Life Sciences Center, Vilnius University, Vilnius, Lithuania

Cardiovascular Institute, Stanford University, Palo Alto, CA, USA

Division of Psychiatry and Behavioral Medicine, College of Human Medicine, Michigan State University, Grand Rapids, MI, USA

L.L.M. contributed to experimental design, computational analyses, immunohistochemistry experiments and cell culture study. B.A.K. performed neuronal nuclei isolations and the bisulfite

padlock probe sequencing of human patient samples, and was involved in the immunohistochemistry and behavioral analyses in mice. E.E. performed the TET2 mRNA analysis in patient samples and

RNA isolation of mouse samples, and contributed to the immunohistochemistry experiments. P.L. was involved in the Hi-C analysis and experimental design of computational approaches. W.C.

performed the bisulfite padlock probe sequencing of the cell culture study. K.X.L. contributed to the immunohistochemistry analysis. M.W. and S.J. were involved in the flow sorting of

neuronal nuclei. J.G. contributed to the hMEDIP analysis. G.A.C. contributed to experimental design. J.M. and X.W. contributed to the analysis of mouse models. V.L. was involved with study

design and overseeing the experiments. The manuscript was written by V.L. and L.L.M. and commented on by all authors.

Peer review information Nature Neuroscience thanks Schahram Akbarian and Tiago Outeiro for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

a, Dendrogram showing unsupervised clustering of samples based on cytosine sites. DNA methylation at top 10,000 most variable cytosine sites. Clustering of corresponding technical (nuclei

sorting and/or library preparation replicated for select DNA samples) and sequencing replicates (sequencing replicated across lanes/flow cells for select libraries) are shown in pink and

purple, respectively. Pearson’s correlation of technical replicates is 0.972 ± 0.0012 s.e.m. and of sequencing replicates is 0.977 ± 0.0010 s.e.m., supporting a high technical

reproducibility. b, The proportion of each CpH subtype for the differentially methylated cytosine sites in PD neurons (significant: orange) as compared to the total number of cytosine sites

profiled (background: gray). CpH context for cytosine sites in enhancers in this study was similar to that of previous whole genome studies in neurons24. c, The proportion of differentially

modified cytosine sites in PD located within an enhancer or promoter. Differentially modified cytosines in neurons of PD patients relative to controls were identified by logistic regression

after adjusting for age, sex, postmortem interval, and neuronal subtypes (n = 57 PD, 48 controls; q