The effect of bone morphogenetic protein-7 on the expression of type I inositol 1,4,5-trisphosphate receptor in G-292 osteosarcoma cells and primary osteoblast cultures.
Bone morphogenetic protein-7 (BMP-7) affects differentiation of preosteoblasts enabling the resultant cells to respond optimally to acutely acting regulators. As the phosphoinositide cascade and, particularly, the calcium-mobilizing inositol 1,4,5-trisphosphate (InsP3) receptor are integral to stimulus-secretion coupling in osteoblasts, the hypothesis that BMP-7 affects InsP3 receptor expression was examined in the G-292 human osteosarcoma cell line and in primary cultures of human osteoblasts. G-292 osteosarcoma cells were found to be a valid experimental model for primary human osteoblasts, expressing osteoblastic mRNAs encoding osteocalcin, bone sialoprotein, alkaline phosphatase, alpha1-collagen, epidermal growth-factor receptor, and BMP type II receptor. When cultured long term in the presence of ascorbic acid and beta-glycerophosphate, G-292 cells underwent further osteoblastic differentiation, forming nodules and exhibiting restricted mineralization. G-292 cells responded to BMP-7 with an increase in InsP3 receptor density. Ligand-binding studies established that BMP-7 (50 ng/ml) treatment of G-292 cells increased InsP3 receptor density 2.4-fold with no apparent change in affinity. Immunoblot analysis with antibodies specific for type I, type II, and type III InsP3 receptors revealed that BMP-7 (50 ng/ml) treatment resulted in a specific increase (206+/-8%) in the type I receptor. Reverse transcription-polymerase chain reaction and Northern blot analyses of G-292 and primary human osteoblasts confirmed an increase in type I InsP3 receptor mRNA upon BMP-7 treatment. These results demonstrate that G-292 cells respond to BMP-7 with an increase InsP3 receptor density, consistent with the enhanced capacity of these cells to respond to Ca2+-mobilizing secretory hormones during osteoblast differentiation.
Bradford
P G
PG
Department of Pharmacology and Toxicology, School of Medicine and Biomedical Sciences, State University of New York at Buffalo, 14214-3000, USA. pgb@acsu.buffalo.edu
Maglich
J M
JM
Ponticelli
A S
AS
Kirkwood
K L
KL
D000469:Alkaline Phosphatase / Q000235:genetics; D001205:Ascorbic Acid / Q000494:pharmacology; D015152:Blotting, Northern; D055419:Bone Morphogenetic Protein 7; D052004:Bone Morphogenetic Protein Receptors; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology*; D002113:Calcification, Physiologic; D015220:Calcium Channels / Q000187:drug effects* / Q000235:genetics; D002454:Cell Differentiation; D002478:Cells, Cultured; D003094:Collagen / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics; D005786:Gene Expression Regulation; D015972:Gene Expression Regulation, Neoplastic; D005994:Glycerophosphates / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D053496:Inositol 1,4,5-Trisphosphate Receptors; D058574:Integrin-Binding Sialoprotein; D010006:Osteoblasts / Q000187:drug effects* / Q000378:metabolism; D015675:Osteocalcin / Q000235:genetics; D012516:Osteosarcoma / Q000235:genetics / Q000473:pathology*; D010716:Phosphatidylinositols / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D011956:Receptors, Cell Surface / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000187:drug effects* / Q000235:genetics; D017978:Receptors, Growth Factor; D012795:Sialoglycoproteins / Q000235:genetics; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
C525819:BMP7 protein, human; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D015220:Calcium Channels; D005994:Glycerophosphates; C546871:IBSP protein, human; C506926:ITPR1 protein, human; D053496:Inositol 1,4,5-Trisphosphate Receptors; D058574:Integrin-Binding Sialoprotein; D010716:Phosphatidylinositols; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D017978:Receptors, Growth Factor; D012795:Sialoglycoproteins; D016212:Transforming Growth Factor beta; D015675:Osteocalcin; D015544:Inositol 1,4,5-Trisphosphate; D003094:Collagen; D066246:ErbB Receptors; D052004:Bone Morphogenetic Protein Receptors; D000469:Alkaline Phosphatase; D001205:Ascorbic Acid; C031463:beta-glycerophosphoric acid