Meeting Abstract
From humans and amphibians to invertebrates, there are many instances of polyploidy of cells with more than 2 copies of a genome per cell. The opisthobranch mollusc, Aplysia californica has endopolyploidy, where central neurons are highly polyploid, but all peripheral tissues are diploid. In fact, Aplysia neurons, some of the largest in the animal kingdom, can have over 100,000 copies of the genome in a single cell. These repeatedly identifiable neurons allowed us to perform single-cell epigenomic profiling and compare the role of DNA methylation in regulating gene expression in polyploid versus diploid cells. We isolated single polyploid neurons (R2, LPL1, MCC) and peripheral diploid tissues (salivary gland and heart) from mature Aplysia that were matched for size and age. We then quantified both genomic DNA copy number in the polyploidy cells and also bisulfite treated gDNA to determine whether each copy of the genome had the same methylation pattern for the same genes. We aimed to test whether multiple genome copies in polyploid neurons are methylated in the same way as diploid cells. Quantal-style analysis of all individual methylation sites implied that polyploidy genome copies are methylated in the same way. Next, to characterize genomic basis of neuronal identify, we have complemented methylome profiling by RNA-seq profiling from the very same identified neurons focusing on serotonergic and cholinergic neurons. As a result, we identified several potential regulators of serotoninergic phenotype and validated expression of these transcription factor candidates using in situ hybridization. This analysis provides the first unbiased view of the genomic regulatory machinery in identified neurons in neural circuits controlling stereotyped and learned behaviors. Supported by NSF, NIH and NASA.
