Figure 6.

Inhibitory effect of Tat-P on TAT activated LTR-Luciferase activity and specific TAR RNA binding by Tat-P. Panel A. The TAT activated LTR-luciferase assay: 239T cells were co-transfected with the pLTR-luc and pCMV-TAT plasmids, and Tat-P and Con-P1 peptides (200 μM, 100 μM, 50 μM) were added to the cells 6 hours after transfection. The conditioned media were exchanged with fresh media containing the same amounts of peptides after 12 hours. The cells were harvested 6 hours later and processed by luciferase assay. The inhibition rates were expressed as mean ± SE. Panel B. The RNA binding assay: 0.25 nmol of TAR RNA was incubated with Tat-P or control peptides (Con-P1 and Con-P2) at indicated Peptide: TAR RNA molar ratio in a total 10 ul of reaction mixture for 15 minutes on ice. Free RNAs and peptide-RNA complexes were resolved by electrophoresis at 25°C on 15% polyacrylamide gels, and imaged using a fluorescent-based EMSA kit.