Figure 5. VSMCs transcription factor gene expressions were negatively and transcriptionally regulated by QKI.

A, B. mRNA levels of SRF, MEF2c and myocardin were significantly down-regulated by QKI over-expression, but up-regulated by QKI suppression. Total RNAs were harvested as described in Figure 4A and 4B. C, D. QKI regulated the gene promoter activities of VSMCs markers and transcription factors. Day 2~3 differentiating ESCs were transfected with luciferase reporter plasmids pGL3-SMA-Luc, pGL3-SM22-Luc, pGL3-SRF-Luc, pGL3-MEF2c-Luc or pGL3-Myocd-Luc (0.15μg/2.5×10⁴ cells) together with GFP-QKI or GFP (0.2μg/2.5×10⁴ cells). Luciferase activity assays were detected 48 hours after transfection. The data presented here are mean±S.E.M. of three to six independent experiments. *P<0.05 (vs. control). E. QKI binds directly to the promoter regions of MEF2c and myocardin genes. ChIP assays were performed using antibodies against QKI or normal rabbit IgG, respectively, as described in the Methods section. PCR amplifications of the specific promoter regions of MEF2c or Myocd gene. The data presented here are mean±S.E.M. of four independent experiments. *P<0.05, #P<0.05 (vs. control).