TYROSINE-HYDROXYLASE IN BOVINE ADRENAL CHROMAFFIN CELLS - ANGIOTENSIN-II STIMULATED ACTIVITY AND PHOSPHORYLATION OF SER(19), SER(31), AND SER(40)

The aim of this study was to determine the effect of angiotensin II (A ll) on tyrosine hydroxylase (TOH) activity and phosphorylation in bovi ne adrenal chromaffin cells (BACCs), We report here that stimulation o f BACCs with All (100 nM) produced a significant increase in both TOH activity and phosphorylation over a period of 10 min. The increase in TOH activity was receptor-mediated. Tryptic phosphopeptide analysis by HPLC revealed that All stimulated an increase in phosphorylation of t hree sites on TOH, Ser(19), Ser(31), and Ser(40), with the largest inc rease being observed for Ser(31) phosphorylation. Pretreatment of the cells with the protein kinase C inhibitor Ro 31-8220 (10 mu M, 15 min) did not affect TOH activity or phosphorylation produced by All. The i nhibitor also did not affect the TOH activity or Ser(40) phosphorylati on produced by forskolin (10 mu M, 10 min). In contrast, Ro 31-8220 fu lly inhibited the TOH activation as well as Ser(31) and Ser(40) phosph orylation of TOH produced by phorbol 12,13-dibutyrate (500 nM, 10 min) . Removal of extracellular Ca2+ from the incubation medium inhibited t he All-induced TOH activity by 50% and significantly blocked Ser(19) a nd Ser(31) phosphorylation but did not affect Ser(40) phosphorylation in response to All. these results indicate that All activates a comple x and perhaps novel signaling pathway leading to the phosphorylation a nd activation of TOH, The TOH activation by All appears to be partiall y independent of Ser(40) phosphorylation, suggesting a potentially imp ortant role for Ser(31) phosphorylation.