Figure 5
PLCβ 2/3 is required for Gβγ dimer-induced PKD activation. (A), Expression of PLCβ2 and PLCβ3 in HEK293 and HeLa cells was determined with their respective anti-sera. HEK293 cells were transfected with pcDNA3 and Gβγ dimers with or without PLCβ3 (B) or PLCβ2 (C). PKD activation was detected by specific phospho-antibodies. Results shown are the mean ± S.E.M. of at least three independent experiments. (D) HEK293 cells were transfected with pcDNA3, PLCβ2, Gβγ with or without PLCβ2. Transfectants were lysed and the extracts analyzed by PLC assay or SDS-PAGE together with Western blot using antibodies against phosphorylated PKD1. Band intensity was quantified by Image J software (National Institute of Health, Bethesda, MD, USA) and depicted in graphical form, and presented as a fold-induction of the pcDNA3 control. Results shown are the mean ± S.E.M. of at least three independent experiments. (E) HeLa cells were transiently transfected with HA-tagged PKD1 together with vector control or Gβγ dimers. The expression of PKD1, FLAG-Gβ, HA-Gγ and endogenous PLCβ3 were detected with their specific antibodies.