Roles of DNA hydroxymethylation versus DNA formylation and carboxylation in neural stem cells [RNA-seq]

TET enzymes are highly expressed in neural stem cells (NSCs). They oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC) in a step-wise manner. In this study we have generated NSCs that only lack the Tet formylation and carboxylation activities (Tet-TFoCa) and have compared their multipotency, transcriptome and epigenome to NSCs that lacked all three Tet activities (Tet-TMut). Our findings reveal unique roles for DNA hydroxymethylation vs DNA formylation and carboxylation in multipotency of NSCs. Overall design: RNA was extracted from neural stem cells at passage one from three lines of wild type and 2 lines of Tet-FoCa and 2 lines of Tet-TMut NSCs using Omega EZNA Total RNA kit. Library preparation and RNAseq were performed by Novogene (https://en.novogene.com/) on a Illumina Novoseq 6000 platform and 20-30 million reads were generated per sample. Details of RNAseq procedure and data analyses are described in detail in the methods section of the manuscript.