It is well established that the phosphorylation of tyrosine hydroxylase (TH) at Ser40 is critical in regulating the catalytic activity of the enzyme. However, the influence of the phosphorylation of Ser40 on the intracellular stability of TH protein has not been investigated. This study was performed to estimate such a possibility. Although the treatment of rat pheochromocytoma cell line PC-12 cells with forskolin increased the amount of TH phosphorylated at Ser40 in the cells, it did not affect the total amount of TH in the cells. Next, human TH type 1 (hTH1) of wild-type and a mutant missing the first 52 amino acid residues were expressed as histidine-tagged forms in PC-12 cells, and then the cells were treated with forskolin. However, the phosphorylation of hTH1 at Ser40 did not affect the amount of the wild-type hTH1 protein present in PC-12 cells. Finally, wild-type and a mutant Ser40 of which was replaced by Asp (S40D, a mimic of TH phosphorylated at Ser40) were expressed in PC-12 cells as histidine-tagged forms or untagged forms. Neither histidine-tagged nor untagged forms showed any difference in their amounts of wild-type hTH1 and S40D hTH1 present in the cells. Collectively, these results indicate the fact that the phosphorylation of Ser40 does not affect the stability of TH protein in PC-12 cells. However, we still do not deny the possibility that the successive phosphorylation of Ser19 and Ser40 may affect the intracellular stability of TH protein. [J Physiol Sci. 2006;56 Suppl:S211]
