Parathyroid hormone induces receptor activity modifying protein-3 (RAMP3) expression primarily via 3′,5′-cyclic adenosine monophosphate signaling in osteoblasts

E. Phelps, O. Bezouglaia, S. Tetradis, J. M. Nervina

Research output: Contribution to journalArticle

Abstract

Parathyroid hormone (PTH) has significant anabolic and catabolic effects on bone. We hypothesize that PTH-induced primary response genes are important determinants of osteoblast function. PTH induces osteoblastic gene expression through PTHR1, a heptahelical receptor that triggers cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA), protein kinase C (PKC), and calcium signaling. By using representational difference analysis we found that receptor activity modifying protein-3 (RAMP3) is a PTH-induced primary response gene in osteoblastic cells. RAMP3 is a coactivator that directs calcitonin receptor (CTR) and CTR-like receptor (CRLR) glycosylation, trafficking, and ligand-binding specificity. Our purpose was to characterize PTH-induced RAMP3 messenger ribonucleic acid (mRNA) levels in primary mouse osteoblasts (MOBs) and to determine which signaling pathway mediates this effect. 10 nM PTH maximally induced RAMP3 mRNA levels in MOBs at 4 hours. Protein synthesis inhibition with 3 μg/mL cycloheximide did not affect PTH-induced RAMP3 mRNA levels. Selective activation of cAMP-PKA signaling with, 10 μM forskolin (FSK) and PKC signaling with 1 μM phorbol 12-myristate 13-acetate (PMA) significantly increased RAMP3 mRNA levels, whereas 1 μM ionomycin (a calcium ionophore) had no effect. Pretreatment with 30 μM H89, a PKA inhibitor, significantly blocked PTH- and FSK-induced RAMP3 mRNA levels. Pretreatment with 1 μM PMA, which depletes PKC, had no effect on PTH- and FSK-induced RAMP3 mRNA levels but blocked PMA-induced RAMP3 mRNA levels. 100 nM PTH (3-34), which activates PKC and calcium but not PKA, had no effect on RAMP3 mRNA levels. These findings indicate that RAMP3 is a PTH-induced primary response gene in primary MOBs and that PTH regulates RAMP3 gene expression primarily through the cAMP-PKA pathway.

Original languageEnglish (US)
Pages (from-to)96-103
Number of pages8
JournalCalcified Tissue International
Volume77
Issue number2
DOIs
StatePublished - Aug 2005

Keywords

  • Osteoblasts
  • PTH
  • Parathyroid hormone
  • RAMP3

ASJC Scopus subject areas

  • Endocrinology, Diabetes and Metabolism
  • Orthopedics and Sports Medicine
  • Endocrinology

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