FIG. 5.
Downregulation of PKC δ elevates PLD activity in EGFR cells. (a) Parental 3Y1 and EGFR cells were treated with EGF (100 ng/ml, 5 min) and/or TPA (400 nM, 24 h) in the presence of 1% butanol, and PLD activity was determined by examining the levels of the PLD-generated transphosphatidylation product phosphatidylbutanol as described in Materials and Methods. The relative PLD activity was normalized to the PLD activity in the untreated 3Y1 cells. Error bars represent the standard deviations for two independent experiments performed in duplicate. (b) The effects of the PKC α- and δ-specific inhibitors Go6976 (0.5 μM) and rottlerin (15 μM) on EGF-induced PLD activity were determined as for panel a. The relative PLD activity was normalized to the PLD activity in the untreated EGFR cells. (c) EGFR cells expressing dominant negative (DN) mutants of PKC α and δ were examined for the effect on PLD activity in the presence and absence of EGF as shown. The relative PLD activity was normalized to the PLD activity in the untreated EGFR cells. Error bars represent the standard deviations for two independent experiments performed in duplicate, where duplicates varied by less than 10%.