Protein Kinase-A and the Effects of Parathyroid Hormone on Phosphate Uptake in Opossum Kidney Cells

Current evidence indicates that signal transduction after receptor binding of PTH involves the stimulation of adenylate cyclase as well as stimulation of phosphoinositide metabolism. Recent studies, showing that PTH alters phosphate transport in opossum kidney cells at concentrations which do not increase cAMP production and that activators of protein kinase-C also alter phosphate transport, have led to the suggestion that there is a dual mechanism for the regulation of phosphate transport by PTH, namely, protein kinase-C at physiological levels of PTH and cAMP at higher levels of PTH. The present studies were designed to evaluate the relationship between cAMP-dependent protein kinase (PK-A), a more sensitive indicator of alterations in cAMP metabolism than measurements of total cellular cAMP, and phosphate transport in opossum kidney cells, in response to bovine (b)PTH 1-34 and [Nle8,Nle18,Tyr34]bPTH 3-34 amide. While bPTH 1-34 markedly stimulated cAMP accumulation (half-maximal stimulation between 1 and 10 nM), PTH 3-34 analog did not. Phosphate transport was inhibited in a dose-dependent manner by bPTH 1-34, with half-maximal effect occurring between 0.1 and 1 nM. [Nle8,Nle18,Tyr34]bPTH 3-34 amide also altered phosphate transport, although this peptide was 3 orders of magnitude less potent than bPTH 1-34. PK-A activity increased in response to bPTH 1-34 and correlated closely with the effects of PTH on phosphate transport. [Nle8,Nle18,Tyr34]bPTH 3-34 amide, which did not appear to increase cAMP, also resulted in a significant increase in the activity of PK-A. Studies of inhibition of cAMP accumulation using 2',5'-dideoxyadenosine demonstrated that while this agent markedly inhibited the accumulation of cAMP in response to PTH, the effects of PTH on phosphate transport were not altered. However, in spite of the reduction in cAMP the activation of PK-A was similar to control. These data indicate that the effects of PTH peptides on phosphate transport are more closely related to changes in the activity of PK-A than to levels of total cAMP. Activation of PK-A in response to PTH is demonstrable at the lowest doses of PTH that alter phosphate transport.