FIG. 6.

The Slap G₁ block involves the C terminus. (A) Fyn overcomes the G₁ block induced by SlapΔC but not by SlapΔSH3. Quiescent NIH 3T3 cells seeded onto coverslips were microinjected with the SlapΔC or SlapΔSH3 constructs alone or with Fyn and stimulated or not with PDGF as shown. The percentage of injected cells that incorporated BrdU for each coverslip was scored as in the legend to Fig. 4A. (B) Fyn does not override the G₁ block induced by the Src/SlapC chimeric adapter. FynΔK or the Src/SlapC adapter constructs were microinjected into seeded quiescent NIH 3T3 cells in the presence or absence of Fyn- or Myc-encoding vectors and stimulated or not with PDGF. Cells were then fixed and processed for immunofluorescence as described in Materials and Methods. The percentage of injected cells that incorporated BrdU for each coverslip was calculated as in the legend to Fig. 4A. (C) Levels of ectopic Slap, SlapΔC, Src, Src527F, and Src/SlapC proteins. Constructs encoding the indicated proteins were transiently transfected into Cos7 cells, and ectopic protein levels were analyzed by Western blotting (wb) of the total cell lysate (10%) and after immunoprecipitation (ip) with the indicated antibodies. The positions of Slap, SlapΔC, Src, Src/SlapC, Ig heavy chain (IgH), and Ig light chain (IgL) are also indicated.