The co-mitogenic combination of transforming growth factor β1 and bombesin protects protein kinase C-δ from late-phase down-regulation, despite synergy in diacylglycerol accumulation

Bombesin induces the down-regulation of protein kinase C-delta (PKC-delta) and PKC-epsilon in Swiss 3T3 cells. Simultaneous addition of transforming growth factor beta 1 (TGF beta 1) selectively blocks PKC-delta down-regulation at mid-S-phase, whereas PKC-epsilon levels continue to decline. Northern blot analysis shows that PKC-epsilon levels could be controlled in part at the level of transcription; PKC-delta mRNA levels remained constant at these later times. Bombesin induces a sustained elevation of some species of diacylglycerol (DAG), consistent with the observed loss of PKC-delta and PKC-epsilon. Interestingly, the combination of bombesin and TGF-beta 1 produces an even greater DAG response. Flow cytometric analysis demonstrates that bombesin induces only 15% of the cells to enter the cell cycle, in contrast to the combination of TGF beta 1 plus bombesin which induces 75-80% of the cells to progress through the cycle. The protection of PKC-delta from down-regulation under conditions of sustained DAG elevation correlates with the mitogenic response and implies that the down-regulation process itself is regulated. Consistent with this, it is demonstrated that bombesin plus TGF beta 1 protects PKC-delta from phorbol ester-induced down-regulation.