Promoter-dependent and -independent activation of insulin-like growth factor binding protein-5 gene expression by prostaglandin E2 in primary rat osteoblasts

Thomas L. McCarthy, Sandra Casinghino, Donald W. Mittanck, Chang Hua Ji, Michael Centrella, Peter Rotwein

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Insulin-like growth factor (IGF) action is mediated by high affinity cell surface IGF receptors and modulated by a family of secreted IGF binding proteins (IGFBPs). IGFBP-5, the most conserved of six IGFBPs characterized to date, uniquely potentiates the anabolic actions of IGF-I for skeletal cells. In osteoblasts, IGFBP-5 production is stimulated by prostaglandin E2 (PGE2), a local factor that mediates certain effects induced by parathyroid hormone, cytokines such as interleukin-1 and transforming growth factor-β, and mechanical strain. In this study, we show that transcriptional and post- transcriptional events initiated by PGE2 collaborate to enhance IGFBP-5 gene expression in primary fetal rat osteoblast cultures. PGE2 treatment stimulated up to a 7-fold rise in steady-state levels of IGFBP-5 mRNA throughout 32 h of incubation. Analysis of nascent IGFBP-5 mRNA suggested that PGE2 had only a modest stimulatory effect on IGFBP-5 gene transcription, and transient transfection studies with IGFBP-5 promoter- reporter genes confirmed that PGE2 enhanced promoter activity by ~2-fold. Similar stimulatory effects were seen with forskolin. A DNA fragment with only 51 base pairs of the 5'-flanking sequence retained hormonal responsiveness, which may be mediated by a binding site for transcription factor AP-2 located at positions -44 to -36 in the proximal IGFBP-5 promoter. Incubation of osteoblasts with the mRNA transcriptional inhibitor 5,6- dichloro-1-β-D-ribofuranosylbenzimidazole demonstrated that PGE2 enhanced IGFBP-5 mRNA stability by 2-fold, increasing the t( 1/2 ) from 9 to 18 h. The effects of PGE2 on steady-state IGFBP-5 transcripts were abrogated by preincubating cells with cycloheximide, indicating that the effects of PGE2 on both gene transcription and mRNA stability required ongoing protein synthesis. Therefore, both promoter-dependent and -independent pathways converge to enhance IGFBP-5 gene expression in response to PGE2 in osteoblasts.

Original languageEnglish (US)
Pages (from-to)6666-6671
Number of pages6
JournalJournal of Biological Chemistry
Volume271
Issue number12
DOIs
StatePublished - Mar 22 1996
Externally publishedYes

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Insulin-Like Growth Factor Binding Protein 5
Osteoblasts
Dinoprostone
Gene expression
Rats
Chemical activation
Gene Expression
Messenger RNA
Insulin-Like Growth Factor Binding Proteins
Genes
RNA Stability
Transcription
Transcription Factor AP-2
Somatomedin Receptors
5' Flanking Region
Transforming Growth Factors
Somatomedins
Colforsin
Cycloheximide
Parathyroid Hormone

ASJC Scopus subject areas

  • Biochemistry

Cite this

Promoter-dependent and -independent activation of insulin-like growth factor binding protein-5 gene expression by prostaglandin E2 in primary rat osteoblasts. / McCarthy, Thomas L.; Casinghino, Sandra; Mittanck, Donald W.; Ji, Chang Hua; Centrella, Michael; Rotwein, Peter.

In: Journal of Biological Chemistry, Vol. 271, No. 12, 22.03.1996, p. 6666-6671.

Research output: Contribution to journalArticle

McCarthy, Thomas L. ; Casinghino, Sandra ; Mittanck, Donald W. ; Ji, Chang Hua ; Centrella, Michael ; Rotwein, Peter. / Promoter-dependent and -independent activation of insulin-like growth factor binding protein-5 gene expression by prostaglandin E2 in primary rat osteoblasts. In: Journal of Biological Chemistry. 1996 ; Vol. 271, No. 12. pp. 6666-6671.
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