Abstract
Parallel single-cell sequencing protocols represent powerful methods for investigating regulatory relationships, including epigenome-transcriptome interactions. Here, we report a single-cell method for parallel chromatin accessibility, DNA methylation and transcriptome profiling. scNMT-seq (single-cell nucleosome, methylation and transcription sequencing) uses a GpC methyltransferase to label open chromatin followed by bisulfite and RNA sequencing. We validate scNMT-seq by applying it to differentiating mouse embryonic stem cells, finding links between all three molecular layers and revealing dynamic coupling between epigenomic layers during differentiation.
Highlights
Parallel single-cell sequencing protocols represent powerful methods for investigating regulatory relationships, including epigenome-transcriptome interactions
It is well known that DNA methylation and other epigenomic layers, including chromatin accessibility, do not act independently of one another[8]
To measure chromatin accessibility together with DNA methylation, we adapted Nucleosome Occupancy and Methylation sequencing (NOMe-seq)[11], where a methyltransferase is used to label accessible DNA prior to bisulfite sequencing (BS-seq), which distinguishes between the two epigenetic states
Summary
Parallel single-cell sequencing protocols represent powerful methods for investigating regulatory relationships, including epigenome-transcriptome interactions. No method that combines RNA-seq with chromatin accessibility profiling in the same cells (with or without DNA methylation) has been reported to-date, which is critical for studying interactions between the epigenome and the transcriptome. A control, we included cells which did not receive enzyme treatment (scM&T-seq controls) and these cells showed universally low GpC methylation levels (~2%), with no enrichment at regulatory regions, indicating that the accessibility data are not affected by endogenous GpC methylation (Supplementary Fig. 3).
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