The purpose of this study was to identify the role of phospholipase D (PLD) isozymes in Bcl-2 expression. Overexpression of PLD1 or PLD2 increased Bcl-2 expression and phosphatidic acid (PA), the product of PLDs, also upregulated Bcl-2 expression. Treatment with PA activated the phospholipase A(2) (PLA(2))/G(i)/ERK1/2, RhoA/Rho-associated kinase (ROCK)/p38 MAPK, and Rac1/p38 MAPK pathways. PA-induced phosphorylation of ERK1/2 was attenuated by a PLA(2) inhibitor (mepacrine) and, a G(i) protein inhibitor (pertussis toxin, PTX). On the other hand, p38 MAPK phosphorylation was attenuated by a dominant negative Rac1 and a specific Rho-kinase inhibitor (Y-27632). These results suggest that PLA(2)/G(i) acts at the upstream of ERK1/2, while Rac1 and RhoA/ROCK act upstream of p38 MAPK. We next, tried to determine which transcription factor is involved in PLD-related Bcl-2 expression. When signal transducer and activator of transcription 3 (STAT3) activity was blocked by a STAT3 specific siRNA, PA-induced Bcl-2 expression was remarkably decreased, suggesting that STAT3 is an essential transcription factor linking PLD to Bcl-2 upregulation. Taken together, these findings indicate that PLD acts as an important regulator in Bcl-2 expression by activating STAT3 involving the phosphorylation of Ser727 through the PLA(2)/G(i)/ERK1/2, RhoA/ROCK/p38 MAPK, and Rac1/p38 MAPK pathways.
