Figure 3.

dfTAT-mediated delivery does not significantly affect cell proliferation and transcription. (a) Proliferation assay. HeLa, Neuro-2a and HDF cells were incubated with 5 μM dfTAT for 1 h or left untreated. Proliferation was assessed using a MTT assay (150,000 cells/experiment, experiments in triplicates, mean and standard deviations represented). (b) Microscopy imaging showing that cells containing cytosolic dfTAT divide. HeLa cells were incubated with 5 μM dfTAT for 1 h, washed and imaged in a time-lapse experiment. Scale bars, 10 μm (c) Whole-genome microarray analysis of HDF cells treated with dfTAT. HDF cells were treated with 5 μM dfTAT for 1h and transcriptome analysis was performed immediately, 1h, or 24h after dfTAT treatment. The plot displays microarray intensity values of treated vs. untreated (same incubation steps but without peptide) samples. The red lines represent 2-fold intensity change cut-offs and transcripts up or down-regulated above these cut-offs are circled for clarity. (d) Microscopy imaging showing that dfTAT-mediated endosomal escape can be repeated. HeLa cells were co-incubated with dfTAT (5 μM) and DEAC-K9 (5 μM) for 1 h (step1) (images not shown). After washing, dfTAT (5 μM) and SNAP-Surface 488 (5 μM) were co-incubated in the absence (top panel) or presence (bottom panel) of bafilomycin (200 nM) (step 3). Scale bars, 10 μm.