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Parathyroid hormone increases prostaglandin G/H synthase-2 transcription by a cyclic adenosine
S Tetradis1, C C Pilbeam, Y Liu
1Department of Medicine, University of Connecticut Health Center, Farmington 06030, USA.
Abstract:
PTH increased PG synthase-2 transcription in osteoblastic MC3T3-E1 cells at 30 min, as assessed by nuclear run-on assays. To determine the signaling pathways used by PTH, the activity of a construct containing the PG synthase-2 gene between nucleotides -963 and +70 linked to a luciferase reporter was analyzed in stably transfected MC3T3-E1 cells. Agents that activate the cAMP-protein kinase A or protein kinase C pathways increased PG synthase-2 promoter activity. In contrast, the calcium ionophore ionomycin was ineffective. The protein kinase A inhibitor H89 blocked PTH stimulation of PG synthase-2 promoter activity, whereas an overnight pre-incubation with phorbol ester to down-regulate protein kinase C did not. PTH-(3-34), a peptide that has greatly reduced ability to activate the cAMP-protein kinase A pathway, did not increase PG synthase-2 transcription or promoter activity. PTH could induce PG synthase-2 messenger RNA accumulation and PG synthase-2 transcription in the presence of cycloheximide. In addition, PTH-stimulated PG synthase-2 transcription was maintained at a high level at 2 h in the presence of cycloheximide. We conclude that PTH rapidly increases PG synthase-2 transcription in MC3T3-E1 cells, mainly through a cAMP-protein kinase A-mediated pathway without the need for protein synthesis. In contrast, the attenuation of increased PG synthase-2 transcription by PTH requires de novo protein synthesis.
Insights
Parathyroid hormone (PTH) rapidly boosts prostaglandin synthase-2 (PG synthase-2) transcription in osteoblasts via the cAMP-protein kinase A pathway. This process does not require new protein synthesis, but its attenuation does.
Area of Science:
- Molecular Biology
- Cellular Biology
- Biochemistry
Background:
- Parathyroid hormone (PTH) plays a crucial role in bone metabolism.
- Prostaglandin synthase-2 (PG synthase-2) is involved in various cellular processes, including bone remodeling.
Purpose of the Study:
- To elucidate the signaling pathways utilized by PTH to regulate PG synthase-2 transcription in osteoblastic cells.
- To investigate the role of cAMP-protein kinase A and protein kinase C pathways in PTH-mediated PG synthase-2 induction.
Main Methods:
- Utilized nuclear run-on assays to assess PG synthase-2 transcription rates.
- Employed reporter gene assays with a PG synthase-2 promoter construct in stably transfected MC3T3-E1 cells.
- Investigated the effects of various signaling pathway activators, inhibitors, and PTH analogs.
Main Results:
- PTH rapidly increased PG synthase-2 transcription and promoter activity in MC3T3-E1 cells.
- The cAMP-protein kinase A pathway was identified as the primary mediator of PTH's effect.
- PTH-induced PG synthase-2 transcription occurred independently of de novo protein synthesis, but its attenuation required it.
Conclusions:
- PTH stimulates PG synthase-2 transcription in osteoblasts predominantly through the cAMP-protein kinase A pathway.
- The rapid induction of PG synthase-2 by PTH does not necessitate protein synthesis.
- De novo protein synthesis is required for the downregulation of PTH-induced PG synthase-2 transcription.