Epigenetic editing to validate findings from methylome-wide association studies of neuropsychiatric disorders

• Main Author: Chan, Robin F.
• Format: Others
• Published: VCU Scholars Compass 2017
• Subjects: epigenetics; CRISPR; DNA methylation; major depressive disorder; psychiatric; editing; Genetics; Molecular Biology; Molecular Genetics; Neuroscience and Neurobiology; Neurosciences; Psychiatric and Mental Health; Psychiatry
• Online Access: http://scholarscompass.vcu.edu/etd/5003; http://scholarscompass.vcu.edu/cgi/viewcontent.cgi?article=6088&context=etd

DNA methylation is necessary for learning, memory consolidation and has been implicated in a number of neuropsychiatric disorders. Obtaining high quality and comprehensive data for the three common forms of methylation in brain is challenging for methylome-wide association studies (MWAS). To address this we optimized a panel of enrichment methods for screening the brain methylome. Results show that these enrichment techniques approach the coverage and fidelity of the current gold standard bisulfite based techniques. Our MBD-based method can also be used with low amounts of genomic material from limited human biomaterials. Psychiatric disorders have high prevalence and are often chronic making them a leading contributor to disability. Major depressive disorder (MDD) has a lifetime prevalence of ~15% and high recurrence leading to substantial morbidity and costs to society. The underlying biological processes that contribute to MDD are poorly understood. Noting the importance of DNA methylation in neurobiology, we conducted the largest MWAS in human post-mortem brain uncover novel candidate genes and biomarkers associated with MDD. The top result of this MDD MWAS was within the gene ANKS1B. This gene has been implicated in many past genetic studies of psychiatric disorders and has experimental support as a regulator of neurotransmission. Targeted epigenetic editing technologies allow for precise modification of DNA methylation in living cells. However, an appropriate model system is critical to properly interpreting such experiments. An accelerated protocol for differentiating Ntera2 cells into human neurons was developed for this purpose. Ntera2-derived neurons express key neuronal markers and are well suited to use in epigenetic editing experiments. Concurrently, the generation of the reagents necessary for recapitulating the aberrant methylation at ANKS1B linked to MDD was undertaken. Using a modified CRISPR/Cas9 approach demethylating enzyme was directed to target sites to attempt perform editing of DNA methylation. Results indicate that significant but biologically irrelevant changes to methylation at ANSK1B were achieved. The novelty of the technology employed presented challenges to the success of the current work. However, the field of epigenetic editing is advancing rapidly and will remain an attractive method for functional characterization of future MWAS findings and basic neuroscience research.