Combining RNA- and protein-centric approaches to discover new functions of non-coding RNAs during stem cell differentiation
Non-coding RNAs are at the interplay between transcription and post transcription regulation. Long non-coding RNAs, in particular, often exert their function by interacting with proteins. In this study, we applied high throughput methodologies combining RNA-centric and protein-centric approaches to identify RNA-protein interactions involved in transcription regulation in pluripotency.
We started by revealing the protein-bound transcriptome of hESCs, embryoid bodies, and each germ layer. We identified the lncRNAs that changed their binding without correlating changes in expression, as well as those unique to pluripotency, differentiated cells, or each germ layer specifically.
We chose the epigenetic factor DNMT3b to further study protein-RNA interactions in hESCs. We found it interacts with the RNAs of 283 genes related to development and transcription, and enriched in CpG islands. DNMT3b’s RNA targets were simultaneously significantly more methylated at the DNA level than other expressed genes, and more expressed. We also found that DNMT3b binds the same motifs at the RNA and DNA level. These DNMT3b/RNA interactions could modulate DNMT3b’s DNA binding and methylation to regulate the expression of key genes in development.