PKC stimulation increases expression of cholinesterases in R28 cell line

Objective: Previously we have shown a counter-regulation between cholinesterases (ChEs) through both anti-sense Butyrylcholinesterase (BChE) transfection and knock-down by siRNAs in R28 cells. In course of this counter-regulation, the status of the cell growth- and differentiation- related signaling pathways PKC and ERK were also changed. Down-regulation of BChE led to an increased PKC-α expression. Here, we demonstrate that this interaction between ChEs and PKC is reciprocal. Methods: R28 cells were treated with final 10 μM Di-octanoyl glycerol (DOG) and 10 nM siRNAs against BChE. Expression analysis was done by RT-PCR, Western Blot, IHC and activity assays. Results: DOG treatment along with BChE knockdown resulted in increased PKC-α expression, as compared with DOG treatment alone. Change in ERK1 expression was not as profound. In R28 cells, DOG stimulation led to marked rapid increase in both AChE and BChE expression. Conclusion: PKC stimulation during BChE knockdown caused functional, saturated AChE expression. Increased PKC-α expression suggests that PKC-α is not only regulated by its stimulators but also by the absence of BChE. While ChEs are counter-regulated, activation of PKC has both reciprocal and additive outcome through cholinergic pathway. Thus, effects of DOG stimulation expose yet another interaction between calcium dependent PKC-α and ChEs showing that any measure of disclosure on neurodegenerative diseases must not only consider cholinergic control but also monitor the PKC pathway.