ELK3 feedback loop with hsa-miR-155-5p.

<p>(A) ELK3 DNA binding to the BIC (MIR155HG) gene promoter as shown in the ChIP-seq normalised bedgraph and by ChIP-QPCR in comparison to the negative locus at Chr12: 96463102–96463229 (Three separate experiments, error bars represent the S.D. between experiments). (B) miR-155 expression in cells treated with siRNA against ELK3 (Four transfection experiments, error bars represent the S.D.). (C) HUVEC cells treated with miR-155, miR-210 or control miR were analysed for mRNA expression of ELK3, HIF1α and NDUFA4 by RT-QCPR (Three separate transfection experiments, error bars represent S.D., **p<0.01, *p<0.05). (D) Protein expression of ELK3 after treatment with siRNA targeting ELK3, control siRNA, miR-155 mimic, control miRNA or miR-155 inhibitor as described in <b>Table S1 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113050#pone.0113050.s001" target="_blank">File S1</a></b>. The loading control was TBP. Quantification of the films from two independent experiments showed that the levels of Elk3 that remained were 40% for siElk3 compared to control, 70% for hsa-miR-155-5p compared to miR Control, and 130% for hsa-miR-155I compared to miRControl. The p values were 0.007, 0.04 and 0.02, respectively, using a Student t-test. (E) ChIP using anti-ELK3 antibody on cells treated with siELK3, miR-155 mimic or control. ELK3 DNA binding was measured at FOS, EGR1, BAD and the negative locus described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113050#pone-0113050-g001" target="_blank">Figure 1</a>. Percentage DNA binding compared to input was measured by QPCR (Two separate transfection experiments, error bars represent the standard deviation).