Evolutionarily conserved regions in the <i>Vegfr3</i> gene act as transcription enhancers in Ras dose-dependent and Ets-dependent manners.

<p>A. Schematic representation of evolutionarily conserved regions in the mouse <i>Vegfr3</i> gene. HR1 and HR2 were identified and described in a previous study <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051639#pone.0051639-Iljin1" target="_blank">[18]</a>. For details, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051639#pone.0051639.s002" target="_blank">Fig. S2</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051639#pone.0051639.s003" target="_blank">Fig. S3</a>. B. Luciferase assay using the pGL4-Luc2 vector harboring the Avr II-Not I fragment (described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051639#pone-0051639-g004" target="_blank">Fig. 4</a>) and fragments derived from HR 3, 5, 6 or 10 in mLECs. The HR fragments used in this study are described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051639#pone.0051639.s003" target="_blank">Fig. S3</a>. Error bars represent the S.D.; n = 3. ****p<0.001 (vs. the HR3-10 fragment-free vector). C. ChIP assay showing H3 histone acetylation on the <i>Vegfr3</i> gene in control, Ras- and Ets-knockdown mLECs. Error bars represent the S.D.; n = 3. *p<0.05, ****p<0.001 (vs. mLECs transfected with control siRNA). Numerical values of the means are shown as % input for rabbit IgG-immunoprecipitated, sonicated chromatin. Results of PCR for <i>Gapdh</i> are shown for comparison. D. ChIP assay showing Ets1 binding to the evolutionarily conserved regions of the <i>Vegfr3</i> gene in control and Ras-knockdown mLECs. Error bars represent the S.D.; n = 3. *p<0.05, **p<0.01 (vs. mLECs transfected with control siRNA).