Generation of stable cell lines overexpressing the wild-type or catalytically inactive forms of PLD1b or PLD2. (A) Two cloned lines with the highest levels of each of the different forms of PLD were selected for detailed analysis. Total lysates from the cells expressing the wild-type PLD1b (D1/WT), catalytically inactive PLD1b (D1/K860R), wild-type PLD2 (D2/WT), catalytically inactive PLD2 (D2/K758R), or empty vector (Vec-1) were immunoblotted with anti-FLAG Ab, anti-PLD1 Ab, or anti-PLD2 Ab (2.5 × 105 cell equivalents/lane for PLD1 and 105 cell equivalents/lane for PLD2). (B) Overexpressed exogenous PLD1b and PLD2 levels were compared by immunoblotting with anti-FLAG Ab (105 cell equivalents/lane). The numbers at the bottom of the lanes are the normalized densitometric analysis.
