Figure 1

Pmel-1 CD8+ T cell transduction and expression of DN TGFβ RII. A. Linear map of DN TGFβ RII retroviral vector, TM = transmembrane region. B. Representative dot plot of mock or DN transduced Pmel-1. C. Untransduced or DN-transduced Pmel-1 T cells were incubated in 10 ng/ml TGFβ for 0, 0.5, 1, 6, 24 hr prior to protein extraction and analyzed for expression of phosphorylated SMAD3. In the presence of TGFβ, untransduced Pmel-1 T cells demonstrated high expression level of phosphorylated SMAD3 protein at 0.5 and 1 hr whereas DN transduced Pmel-1 T cells did not show p-SMAD3 expression, indicating DN transgene effectively inhibits TGFβ signaling in transduced cells. D. Absolute cell counts of Pmel-1 cells re-activated with IL-2 (50 IU/ml) and gp100 peptide (1 μg/ml) and treated with mouse TGFβ1 (10 ng/ml) for 72 h.