Structure-function relationship of parathyroid hormone: activation of phospholipase-C, protein kinase-A and -C in osteosarcoma cells.
Recent evidence indicates that after PTh interaction with its receptor, both protein kinase-A (PKA) and protein kinase-C (PKC) are activated. To investigate the relationship between PTH structure and protein kinase stimulation, we have analyzed the effects of synthetic PTH fragments on PKA and PKC in the rat osteogenic sarcoma cells, UMR 106-01. Activation of PKA by 10(-7) M bovine (b) PTH-(1-34) was maximal (2.7-fold of control) at 5 min and remained elevated 15 min after hormone exposure. bPTH-(2-34), at equimolar doses, also stimulated PKA, but with a lower potency (1.4-fold of control), whereas propionyl bPTH-(2-34) [pbPTH-(2-34)], bPTH-(3-34), [Tyr34]bPTH-(7-34) amide [bPTH-(7-34)], and bPTH-(30-34) were ineffective. On the other hand, translocation of PKC activity from the cytosol to the membrane after exposure to bPTH-(1-34) was transient, with a peak at 1 min (1.9-fold of control), and returned to basal levels after 5 min. Other fragments, bPTH-(2-34), pbPTH-(2-34), bPTH-(3-34), and bPTH-(7-34), were also active on PKC, with relative potencies of 81%, 67%, 62%, and 51% of bPTH-(1-34), respectively, whereas bPTH-(30-34) was inactive. bPTH-(1-34), bPTH-(2-34), pbPTH-(2-34), and bPTH-(3-34) also induced inositol 1,4,5-trisphosphate production, with a potency order of 1.6-, 1.6-, 1.5-, and 1.6-fold over the control value, respectively, thus indicating activation of phospholipase-C. Neither bPTH-(7-34) nor bPTH-(30-34) caused a statistically significant increase in inositol 1,4,5-trisphosphate production. These results demonstrate that PTH signal transduction through the two different pathways can be dissociated; while activation of the cAMP/PKA system requires amino acids 1 and 2, the phospholipase-C/PKC system is coupled to a longer domain of the hormone's N-terminus.
Fujimori
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A
Division of Bone and Mineral Metabolism, Jewish Hospital of St. Louis, Washington University Medical Center, Missouri 63110.
Cheng
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SL
Avioli
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LV
Civitelli
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R
D000818:Animals; D002118:Calcium / Q000378:metabolism; D004789:Enzyme Activation; D007295:Inositol Phosphates / Q000378:metabolism; D012516:Osteosarcoma / Q000201:enzymology*; D010281:Parathyroid Hormone / Q000494:pharmacology*; D010446:Peptide Fragments / Q000494:pharmacology*; D011493:Protein Kinase C / Q000032:analysis*; D011494:Protein Kinases / Q000032:analysis*; D051381:Rats; D013329:Structure-Activity Relationship; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D010738:Type C Phospholipases / Q000032:analysis*
D007295:Inositol Phosphates; D010281:Parathyroid Hormone; D010446:Peptide Fragments; D011494:Protein Kinases; D011493:Protein Kinase C; D010738:Type C Phospholipases; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium