Figure 5
Activating histone modifications affect transcriptional activation for DHRS4-AS1, DHRS4L2-AS and DHRS4L1-AS.
(A–C) Relative levels of acetyl-H3, H3K4me3 and H3K9me2 histone modifications were determined by ChIP analysis at predicted promoter regions within DHRS4-AS1 and its putative homologous NATs of the DHRS4 gene cluster in HepG2 cells and HL7702 cells. DHRS4-AP refers to the predicted promoter of DHRS4-AS1 and DHRS4L2-AP refers to the predicted promoter regions of putative NAT of DHRS4L2. Putative NAT promoter regions of DHRS4L1 were termed DHRS4L1-AP1, DHRS4L1-AP2, DHRS4L1-AP3 and DHRS4L1-AP4 according to the position at different gene loci (Methods). An unpaired Student’s t test was used to evaluate the differences between DHSRS4-AP and the other promoters. Error bars represent the mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001. (D) H3K27me3 is present at very low levels at the predicted promoter regions within DHRS4-AS1 and its putative homologous NATs in HL7702 and HepG2 cells. In ChIP assays, the PCR bands of α-satellite primers were used as the positive control to biologically validate the IP results. IgG was the negative control to ensure specificity of the ChIP reaction. (E) CpG island methylation status within DHRS4-AS1 and its putative corresponding NAT regions was detected by using BSP analysis in HL7702 and HepG2 cells. Closed circles represent methylated CpG sites and open circles represent unmethylated CpG sites. The relative locations of CpG islands at the putative homologous NATs are shown (lower panel).
