Hormonal control of insulin-like growth factor I gene transcription in human osteoblasts: dual actions of cAMP-dependent protein kinase on CCAAT/enhancer-binding protein delta

Insulin-like growth factor-I (IGF-I) is essential for somatic growth and promotes bone cell replication and differentiation. IGF-I production by rat osteoblasts is stimulated by activation of cAMP-dependent protein kinase (PKA). In this report, we define two interacting PKA-regulated pathways that control IGF-I gene transcription in cultured human osteoblasts. Stimulation of cAMP led to a 12-fold increase in IGF-I mRNA and enhanced IGF-I promoter activity through a DNA response element termed HS3D and the transcription factor CCAAT/enhancer-binding protein delta (C/EBPdelta). Under basal conditions, C/EBPdelta was found in osteoblast nuclei but was transcriptionally silent. Treatment with the PKA inhibitor H-89 caused redistribution of C/EBPdelta to the cytoplasm. After hormone treatment, the catalytic subunit of PKA accumulated in osteoblast nuclei. Inhibition of active PKA with targeted nuclear expression of PKA inhibitor had no effect on the subcellular location of C/EBPdelta but prevented hormone-induced IGF-I gene activation, while cytoplasmic PKA inhibitor additionally caused the removal of C/EBPdelta from the nucleus. These results show that IGF-I gene expression is controlled in human osteoblasts by two PKA-dependent pathways. Cytoplasmic PKA mediates nuclear localization of C/EBPdelta under basal conditions, and nuclear PKA stimulates its transcriptional activity upon hormone treatment. Both mechanisms are indirect, since PKA failed to phosphorylate human C/EBPdelta in vitro.