Deciphering Motor Dysfunction and Microglial Activation in mThy1-a-Synuclein Mice: A Comprehensive Study of Behavioral, Gene Experssion, and Methylation Changes [Enzymatic Methyl-Seq]
BACKGROUND: Synucleinopathy disorders are characterized by aggregates of a-synuclein (a-syn), which engage microglia to elicit a neuroinflammatory response. However, the influence a-syn has on DNA methylation and gene expression patterns for other genes has not been thoroughly explored. Our objective is to determine gene expression and methylation changes in microglia induced by aggregate a-syn. METHODS: Transgenic mThy1-Asyn mice overexpressing human a-syn are a model of synucleinopathy. Microglia from 7 and 10-month-old mice were used to isolate nucleic acids for methylated DNA and RNA-sequencing. a-Syn-induced changes in gene expression and genomic methylation were determined and examined for pathway and functional enrichment. RESULTS: Microglial DNA isolated from our 7-month cohort had 6,712 differentially methylated regions (DMR), while RNA levels demonstrated a change in 1,141 differentially expressed genes (DEGs) between mThy1-Asyn mice and wild-type littermate controls. The 7-month DEGs and DMRs were highly associated with neuroinflammation likely via TLR pathways. We observed 8,531 DMRs and 1,686 DEGs in 10-month mThy1-Asyn mice. These genes were often predicted by Ingenuity Pathway Analysis to demonstrate inhibition of previously activated inflammatory pathways at 7 months. CONCLUSION: We observed significant a-syn-induced methylation and gene expression changes in microglia. Our data suggest that a-syn overexpression initiates microglial activation leading to neuroinflammation and cellular metabolic stresses, which is associated with disease progression. Overall design: We performed transcriptomic and methylomic analysis of murince brain homogenate subjected to IBA1 selection for microglia from mice overexpressing full-length human wild-type a-syn under the murine Thy-1 promoter on the X chromosome. Transcriptomic and methylomic profiles were analyzed for biological meaning using Gene ontology and Ingenuity Pathway Analysis on differentially expressed genes (DEGs) and genes annotated to differentially methylated regions (DMRs). HEM samples are mice overexpressing human alpha-synuclein while WT mice are controls. The corresponding background of these mice is the B6D2F1/J mice (stock# 100006) from The Jackson Laboratory (Bar Harbor, ME). There are five 10-month-old HEM mice and five 10 month wild-type controls. There are six 7-month HEM mice and four 7-month wild-type controls.