Augmentation of Tax-induced activation of HTLV-1 LTR by group I Paks. (A and B) Augmentation of LTR activation in HeLa cells. Cells were transfected with the indicated reporter and expression plasmids. A fixed amount of Tax expression plasmid plus escalating amounts of expression plasmids for Pak1/2/3 (A) or a fix amount of Pak3 expression plasmid plus escalating amounts of Tax plasmid (B) were used. Numbers at the top of the error bars indicate fold activation. (C) Augmentation of TRE activation in HeLa cells. The reporter plasmid pTRE-Luc contains three copies of TRE only. (D) Augmentation of LTR activation in Jurkat cells. (E) RNAi depletion of Pak1 or Pak3 dampened LTR activity in HTLV-1-transformed T cells. MT2 cells were transfected with pre-validated siRNA (100 nM) against Pak1 (siPak1A/B) or Pak3 (siPak3A). Cells were transfected with pLTR-Luc 30 h after siRNA transfection. Proliferation of MT2 cells were unaffected by siPak as measured by MTT assay. *: the difference between groups 6 and 5 is statistically significant (p = 0.0020 by Student's t test). #: p = 0.0017. @: p = 0.0018. (F) Paks have no influence on Tax-induced activation of NFκB in HeLa cells. Dual luciferase assays were performed. Relative luciferase activity (RLA) was calculated as a ratio of firefly luciferase activity recovered from pLTR-Luc or pHIAP-Luc versus that of Renilla luciferase recovered from co-transfected pSV-RLuc. Results represent three independent experiments and standard deviations (SD) are indicated by error bars.