RRC ID 73331
Author Fukuda T, Takekoshi K, Nanmoku T, Ishii K, Isobe K, Kawakami Y.
Title Inhibition of the RhoA/Rho kinase system attenuates catecholamine biosynthesis in PC 12 rat pheochromocytoma cells.
Journal Biochim Biophys Acta
Abstract The small GTPase, RhoA, and its downstream effecter Rho-kinase (ROK) are reported to be involved in various cellular functions, such as myosin light chain phosphorylation during smooth muscle contraction and exocytosis. Indeed, growing evidence suggests that the RhoA/Rho-kinase pathway plays an important role in regulating exocytosis in these cells. However, it is not known whether the RhoA/Rho-kinase pathway has an effect on catecholamine synthesis. Using the rat pheochromocytoma cell line, PC12, we examined the effects of either Rho-kinase inhibitor (Y27632) or RhoA inhibitor (C3 toxin) on nicotine-induced catecholamine biosynthesis. We show that nicotine (10 microM) induces a significant, though transient, increase in RhoA activation in these cells. Treatment with either Y27632 (1 microM) or C3 toxin (10 microg/ml) significantly inhibited the nicotine-induced increase of tyrosine hydroxylase (TH) mRNA and the corresponding enzyme activity. TH catalyzes the rate-limiting step in the biosynthesis of catecholamine. Y27632 significantly inhibited nicotine-induced phosphorylation of TH at Ser40 as well as Ser19, which are known to be phosphorylated by Ca(2+)/calmodulin kinase II. Furthermore, Y27632 (10 microM) as well as C3 toxin (10 microg/ml) significantly inhibited the nicotine-induced increase of TH at the protein level. Thus, we propose that activation of RhoA, and its downstream effecter Rho-kinase, is a prerequisite for catecholamine biosynthesis in PC12 cells. At the concentrations used in our experiments, Y27632 does not affect cAMP/PKA activity or PKC activity, indicating that the inhibitory effect of Y27632 can be attributed to the inhibition of Rho-kinase activity as observed in chromaffin cells. In contrast, neither Y27632 (10 microM) nor C3 toxin (10 microg/ml) significantly altered catecholamine secretion in PC12 cells. In conclusion, we have demonstrated that inhibition of the Rho/Rho-kinase pathway in chromaffin cells lowers TH activity, probably through CaMKII inhibition. By contrast, neither Y27632 nor C3 toxin affect the secretion of catecholamine.
Volume 1726(1)
Pages 28-33
Published 2005-10-30
DOI 10.1016/j.bbagen.2005.08.008
PII S0304-4165(05)00261-8
PMID 16219424
MeSH Amides / pharmacology Analysis of Variance Animals Blotting, Western Botulinum Toxins / pharmacology Catecholamines / biosynthesis* Cyclic AMP / metabolism Cyclic AMP-Dependent Protein Kinases / metabolism DNA Primers Gene Expression Regulation, Enzymologic / drug effects* Glutathione Transferase Intracellular Signaling Peptides and Proteins Nicotine / metabolism PC12 Cells Phosphorylation Polymerase Chain Reaction Protein Kinase C / metabolism Protein Serine-Threonine Kinases / antagonists & inhibitors* Protein Serine-Threonine Kinases / metabolism Pyridines / pharmacology Rats Signal Transduction / drug effects* Tyrosine 3-Monooxygenase / metabolism rho-Associated Kinases rhoA GTP-Binding Protein / antagonists & inhibitors* rhoA GTP-Binding Protein / metabolism
IF 3.411
Human and Animal Cells PC-12(RCB0009)