Characterization of HTLV-1 gene expression in SLB-1 cells stably infected with shRNA lentiviral vectors. (A) Total RNA was isolated from parental SLB-1, stably transduced SLB-1 (V1-V5), and control Jurkat cells and subjected to real-time RT-PCR to quantify Hbz transcripts. Numbers are normalized to 106 copies of hGAPDH. Cells transduced with Hbz-specific shRNA vectors (V2-V4) have a significant 3- to 3.5-fold decrease in Hbz mRNA levels compared with controls (*). (B) Western blot analysis on total cellular lysates from indicated cells shows a significant 3-fold decrease in HBZ protein expression (normalized HBZ densitometry numbers) in cells transduced with Hbz-specific shRNA vectors (V2-V4). Protein reduction correlates directly to mRNA reduction. (C) Quantitation of tax/rex and gag/pol mRNA expression in total cellular RNA by real-time RT-PCR (normalized to 106 hGAPDH) and (D) p19 Gag expression in culture supernatant by ELISA. The data presented in panels A, C, and D are the average values from 3 independent experiments; error bars denote SD. Statistical significance was determined by analysis of variance (ANOVA) followed by Tukey test.
