Synthesis and biological activity of Pdx-1, TAT-Pdx-1, Pdx-1-VP16 and TAT-Pdx-1-VP16 fusion proteins. (a) Schematic structure (top panel) and purity (bottom panel) of Pdx-1, TAT-Pdx-1, Pdx-1-VP16 and TAT-Pdx-1-VP16 fusion proteins. PTDPdx-1 represents protein transduction domain of the Pdx-1 protein, which is naturally present in the four Pdx-1(-VP16) fusion proteins. TAT represents protein transduction domain of the VIH TAT protein. VP16 is the activation domain of Herpes simplex virus I. His represents hexahistidine tag used to purify proteins by His-tag affinity chromatography. Purified proteins were run on a SDS-PAGE gel (8%) stained with Coomassie blue. Molecular weights are 45 kDa for Pdx-1 and TAT-Pdx-1, 60 kDa for Pdx-1-VP16 and TAT-Pdx-1-VP16. (b) Insulin promoter activity after Pdx-1(-VP16) protein transduction. Twelve hours after transient transfection of insulin promoter luciferase plasmid, HepG2 cells were treated with 5 μM Pdx-1(-VP16) fusion proteins for 36 hours. Then luciferase activity was measured. Data are expressed as mean + SEM (Control n = 14, Pdx-1 n = 13, TAT-Pdx-1 n = 7, Pdx-1-VP16 n = 12, TAT-Pdx-1-VP16 n = 19). Insulin promoter activity of controls was arbitrarily set at 1. Statistical comparisons were performed using a non parametric Mann-Withney test, *: p < 0.05, ***: p < 0.001.
Delisle et al. BMC Research Notes 2009 2:3 doi:10.1186/1756-0500-2-3