Unraveling the role of ZBED6 in transcriptional regulation by whole transcriptome analysis after RNAi in mouse myoblasts

ZBED6 is a novel transcription factor unique to placental mammals and has evolved from a domesticated DNA transposon. Here we further characterize the functional significance of ZBED6 based on transcriptome analysis of mouse myoblasts after Zbed6 silencing. ZBED6 appears as an important transcriptional regulator since differential expression of more than 1,000 transcripts was consistently observed after Zbed6 silencing and these changes correlated with increased myotube formation. Up-regulated genes were associated with muscle contractile fibers and muscle organ development, while down-regulated genes were associated with ribosome function. Thirty-four small nucleolar RNAs showed differential expression and all increased in expression after Zbed6 silencing. This is particularly interesting since ZBED6 shows a strong enrichment in the nucleolus. There was an overrepresentation of genes with ZBED6 binding sites among the up-regulated genes after silencing, suggesting that ZBED6 acts as a repressor at many loci. Many genes showed significant down-regulation after Zbed6 silencing, which begs the question of whether ZBED6 acts as an activator at some of these loci or if they all represent secondary effects. The colocalization of histone marks and ZBED6 binding sites defined by a previous ChIP-seq analysis was evaluated. There was a strong association between ZBED6 binding sites and the H3K4me3, H3K4me2 and H3K27ac modifications, which are usually found at active promoters, but no association with the repressive marks H3K27me3 and H3K36me3. We propose that ZBED6 preferentially binds to active promoters and modulates transcriptional activity by a novel mechanism rather than by recruiting repressive histone modifications. C2C12 cells were either exposed to siRNA targeting ZBED6 or negative control siRNAs. Cells in each treatment group were cultured for 48 and 96 hours before RNA was collected. Each treatment and time point was represented by three biological replicates.