FIG. 2.

Existence of a calcium-independent targeting of PKCα. (A) PKCα and -β1 behaved differently when GH3B6 cells were incubated with 2 μM ionomycin. PKCβ1 is activated and accumulated at the entire plasma membrane similar to what is seen in the presence of 100 nM TRH. PKCα was not activated and remained in the cytoplasm. (B) PKCα and -β bearing the triple point mutations D187/246/248N do not translocate in the presence of ionomycin. mut, mutant. (C) D187/246/248N PKCα, unable to bind calcium, was still targeted to the cell-cell contacts upon stimulation, indicating that its translocation can be calcium independent. The same mutations in the PKCβ1 sequence show that PKCβ1 behaved like PKCα when unable to bind calcium: it accumulated at cell-cell contacts. Bar, 5 μm. (D) An example of intracellular calcium modification upon TRH stimulation. Image acquisition was performed on a confocal microscope, with 0.533-s intervals between frames. Calcium monitoring was described previously (48). F/Fmin, fluorescence/minimum fluorescence. (E) Summary of the results as described in the legend to Fig. 1C.