Effects of recombinant TAT-Gab2 fusion proteins on cS5^F-induced cell growth and Akt phosphorylation. (A) Schematic representation of TAT-Akt and TAT-Gab2 constructs. The different cDNAs were introduced into the bacterial expression vector pTAT-HA. Resulting recombinant Akt (wt and dn) and Gab2 (wt and Gab23YF) proteins were fused in their N-terminal part to a 6 ×His-Tag followed by the protein transduction domain (PTD) of the TAT protein and a HA tag sequence. (B) cS5^F BM cells were transduced or not with 100 nM of the different TAT-Gab2 proteins during 48 hours, and the number of living cells was determined daily using the trypan blue dye assay. Results are the mean of 3 independent experiments performed with 2 independent cS5^F BM-cell cultures. (C) GFPv BM cells and cS5^F BM cells were transduced with 100 nM TAT-Gab23YF fusion protein, and growth of the cells was determined daily. Results are the mean of 3 independent experiments. (D) After extensive washes, lysates from transduced cS5^F BM cells were prepared and analyzed by Western blotting with the indicated antibodies. Densitometric analysis was also performed to determine the ratio of P-Akt/Akt in the different samples (bottom).