Matrix GLA protein modulates differentiation induced by bone morphogenetic protein-2 in C3H10T1/2 cells.
Matrix GLA protein (MGP) is ubiquitously expressed with high accumulation in bone and cartilage, where it was found to associate with bone morphogenetic proteins (BMP) during protein purification. To test whether MGP affects BMP-induced differentiation, three sets of experiments were performed. First, pluripotent C3H10T1/2 cells transfected with human MPG (hMGP) or antisense to hMGP (AS-hMGP) were treated with BMP-2. In cells overexpressing hMGP, osteogenic and chondrogenic differentiation was inhibited indicating decreased BMP-2 activity. Conversely, in cells overexpressing AS-hMGP, BMP-2 activity was enhanced. Second, cells were prepared from homozygous and heterozygous MPG-deficient mice aortas. When treated with BMP-2, these cells underwent chondrogenic and osteogenic differentiation, respectively, whereas controls did not. Third, FLAG-tagged hMGP with the same biological effect as native hMGP inhibited BMP-induced differentiation, when exogenously added to culture media. Together, these results suggest that MGP modulates BMP activity. To test whether hMGP fragments would retain the effect of full-length hMGP, three subdomains were overexpressed in C3H10T1/2 cells. In cells expressing the mid-region, alone (amino acids (aa) 35-54) or in combination with the N terminus (aa 1-54) but not the C terminus (aa 35-84), osteogenic differentiation was enhanced and occurred even without added BMP-2. Thus, two subdomains had the opposite effect of full-length hMGP, possibly due to different expression levels or domain characteristics.
Boström
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K
Division of Cardiology, Departments of Medicine and Physiology, UCLA School of Medicine, Los Angeles, California 90095-1679, USA. kbostrom@mednet.ucla.edu
Tsao
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D
Shen
S
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Wang
Y
Y
Demer
L L
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D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D001011:Aorta / Q000378:metabolism; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation; D002460:Cell Line; D003094:Collagen / Q000378:metabolism; D005822:Genetic Vectors / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D008648:Mesoderm / Q000166:cytology; D051379:Mice; D008809:Mice, Inbred C3H; D015675:Osteocalcin / Q000096:biosynthesis* / Q000378:metabolism / Q000502:physiology; D010641:Phenotype; D010766:Phosphorylation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D016212:Transforming Growth Factor beta
C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D015675:Osteocalcin; D003094:Collagen; D000469:Alkaline Phosphatase