Functional genomics analysis in human embryonic stem cells using RNA interference.

Abstract

Human embryonic stem cells (hESCs) have the ability to proliferate indefinitely and remain undifferentiated due to their pluripotent characteristic. However, the mechanisms underlying their ability for self-renewal are still not well characterised. Key transcriptional regulators NANOG, OCT4 and SOX2 make up the well-known core transcriptional regulatory network that maintains hESC pluripotency. In order to discover novel factors that control hESC self-renewal, some genes of potential to influence the activity of NANOG are identified through preliminary screens. With the use of small interfering RNAs (siRNAs) and cDNA clones in expression vectors, effects on NANOG activity from the knockdown or overexpression of BRD4, SIRT1, HEXIM2 and HDAC5 in hESCs were studied. From the results, effects of these genes are better understood. They appeared to have an indirect influence on NANOG activity. Distinctively, BRD4 caused reduced induction of NANOG activity in both knockdown and overexpression studies. This present study will serve as a platform for further investigations into the effects of BRD4, SIRT1, HEXIM2 and HDAC5 on properties of hESC including pluripotency.