Gene expression profiles of depletion of Daxx and HOTAIR

In this study, we found that Daxx may modulate HOTAIR stability, and therefore, regulate H3K27me3 levels. To further examine the possible mechanism and biological consequence, we separately knocked down Daxx and HOTAIR with a lentivirus plasmid constructed with short hairpin RNA and siRNA oligonucleotides, respectively. We then used microarray analysis to identify several candidate genes regulated under the Daxx-HOTAIR axis. In order to knock down Daxx, HEK293T cells were transfected with a lentivirus plasmid constructed with a short hairpin RNA targeted to Daxx or to luciferase as a control. In the HOTAIR experiment, siRNA oligonucleotides were targeted to HOTAIR or to GFP as a control. Both were synthesized by Dharmacon. All transfections were performed by using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instructions. Three independent knockdown experiments were performed with Daxx and HOTAIR. Total RNA was extracted after 3 days to perform microarray analysis.