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. 2009 Sep;11(9):921–933. doi: 10.1593/neo.09636

Figure 2.

Figure 2

LEDGF/p75 transcriptionally activates VEGF-C expression in an STRE-dependent manner. A luciferase reporter gene containing the 5′-flanking region of human VEGF-C gene (Figure 1A, pVEGF-C-Luc) was transiently cotransfected with a control empty vector (pIRES) or with a construct encoding LEDGF/p75 into rat glioma C6 or COS7 cells (A and B, respectively). Luciferase activity under each experimental condition was scaled relative to the activity in control cells (mean ± SD, n = 3). (C and D; upper panel) Western blot analysis carried out using anti-HA antibodies of protein extracted from C6 rat glioma (C) or human H1299 (D) cells stably expressing the rat LEDGF protein tagged to hemagglutinin (HA) epitope (pIRES-LEDGF) after selection with puromycin. (C and D; lower panel) Measurement of luciferase activity in C6 (C) or H2299 (D) cells stably expressing either LEDGF protein (pIRES-LEDGF) or the empty control vector (pIRES) transiently transfected with the pVEGF-C-Luc construct. (E) Wild type (wt) and two mutated (m1 and m2) sequences of the conserved LEDGF/p75 binding sites are presented, and the G-to-A substitutions are indicated in gray. (F) H1299 cells were transiently cotransfected either with intact pVEGF-Cwt-Luc reporter or pVEGF-Cm1-Luc, pVEGF-Cm2-Luc together with LEDGF/p75 expression vector or a control empty vector (+ and - indicate presence and absence of each construct; mean ± SD, n = 3).