Ten-Eleven Translocation Proteins-Mediated Epigenetic Regulation in Alzheimer’s Disease Restricted; Files Only
Armstrong, Matthew (Spring 2023)
Published
Abstract
The characterization of Alzheimer’s disease (AD) is complex and multifactorial with genetic, epigenetic, and environmental factors contributing toward pathogenesis. Barring familial dominantly inherited forms of AD, the interplay between these three factors influences whether an individual develops AD. TET enzymes function as key regulators of DNA methylation and hydroxymethylation profiles, and, in conjunction with their other functions of transcription factor (TF) binding and recruitment, dynamically regulate spatiotemporal gene expression. Prior association and functional studies have identified TET enzymes as important players in the development of various diseases ranging from cancer to neurodevelopmental, neuropsychiatric, and neurodegenerative disorders and diseases. To build upon previous research in the field, here, we investigate the role of TET enzymes in AD. We utilize a multifaceted approach to explore the potential role of TET in human AD pathogenesis, as well as investigate how the functional loss of Tet1 and Tet2 impact AD molecular and pathological phenotypes in the 5xFAD mouse model. Given the association between AD and perturbation DNA methylation profiles, identified through EWAS studies, the EWASplus algorithm was developed to utilize existing array methylation data that covers only 2% of CpG sites in the genome, in conjunction with additional informative epigenetic features, to predict the association of off-array CpG sites with LOAD. Select EWASplus predicted sites were experimentally validated, and the EWAS plus algorithm was further utilized to classify human samples into AD and control groups prior to performing hydroxymethylation and gene expression profiling which revealed DhMRs in DNMT3A, DNMT3L, TET3, and a significant increase in expression of AD genes TREM2 and FCER1G. Genetic variant analysis performed via targeted re-sequencing of TET1, TET2, and TET3 identified individual TET1 missense variants as well as an overall enrichment of TET1 variants associated with EOAD. Lastly, functional analysis performed in the 5xFAD mouse model identified that the loss of Tet1 and Tet2 promote strikingly similar contributions towards AD outcome by increasing AD-associated pathology, disrupting 5hmC landscapes, and increasing the expression of AD-associated genes Trem2, Mpeg1, and C1qa. Altogether, this work characterizes the importance of the methylome, hydroxymethylome, and TET enzymes as mediators of AD pathogenicity.
Table of Contents
Chapter 1 - Diverse and dynamic DNA modifications in brain and diseases. 7
ABSTRACT. 8
INTRODUCTION.. 8
Cytosine Modifications (5mC, 5hmC, 5fC and 5caC) 9
Establishment, maintenance and function of 5mC.. 9
Establishment, maintenance, and function of 5hmC derivatives (5hmC, 5fC, and 5caC) 12
N6-methyladenine. 15
DNA Modifications in Neurodegenerative Disorders. 19
Alzheimer’s disease. 19
Parkinson’s disease. 21
Ataxia-related disorders. 22
Spinocerebellar ataxias. 23
Fragile X-associated tremor/ataxia syndrome. 24
Possible New Roles of DNA Modifications. 25
Modulators of dynamic TF and DNA interaction. 25
Brain genomic mosaicism.. 27
Summary. 28
ACKNOWLEDGEMENTS.. 29
FIGURES.. 30
REFERENCES.. 33
Chapter 2 - A machine learning approach to brain epigenetic analysis reveals kinases associated with Alzheimer’s disease. 44
ABSTRACT. 45
INTRODUCTION.. 45
RESULTS. 47
EWASplus overview.. 47
EWASplus performance compared to methylation array. 48
EWASPlus performance for off-array CpGs. 50
Comparison with a competing method. 51
Experimental validation of EWASplus predictions. 51
EWASplus performance on multiple cohorts. 52
Biological insights into AD.. 53
DISCUSSION.. 54
METHODS. 59
FIGURES.. 69
TABLES.. 74
Supplementary Figures. 78
REFERENCES.. 96
Chapter 3 - Role of TET1-mediated epigenetic modulation in Alzheimer’s disease. 101
SUMMARY. 102
INTRODUCTION.. 103
RESULTS. 106
Tet1 Genetic Variants Are Associated With EOAD.. 106
5hmC and Gene Expression Dysregulation of Key AD and DNA Modification Regulator Genes Are Observed in LOAD.. 106
Loss of Tet1 Increases AD-associated Pathology in 5xFAD Mice. 108
Tet1 Deficiency Alters 5xFAD-associated Epigenetic Landscapes. 110
Partial Loss of Tet1 Modifies 5xFAD-associated Gene Expression. 112
DISCUSSION.. 113
ACKNOWLEDGEMENTS.. 116
DECLARATION OF INTEREST. 116
TABLES.. 117
FIGURES.. 121
Supplementary Figures. 130
METHODS. 135
REFERENCES.. 145
Chapter 4 – Tet2 deficiency exacerbates AD-associated molecular and neuropathology in the 5xFAD mouse model 153
SUMMARY. 154
INTRODUCTION.. 155
RESULTS. 157
Loss of Tet2 Increases AD Physiopathology in 5xFAD Mice. 157
Loss of Tet2 Promotes Dysregulation of 5xFAD-Associated 5hmC Profiles. 158
Partial Loss of Tet2 Disrupts 5xFAD-Associated Gene Expression. 159
Similar, Yet Discrete Effects Caused by the Loss of Tet1 and Tet2 on AD Pathogenesis in 5xFAD Mice. 161
DISCUSSION.. 162
FIGURES.. 166
Supplementary Figures. 178
TABLES.. 181
METHODS. 184
REFERENCES.. 189
Chapter 5 – Summary, Limitations, Future Directions, and Conclusions. 197
SUMMARY. 197
OVERALL LIMITATIONS. 199
FUTURE DIRECTIONS.. 201
CONCLUSIONS.. 202
REFERENCES.. 204
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