Dentin phosphophoryn activates Smad protein signaling through Ca2+-calmodulin-dependent protein kinase II in undifferentiated mesenchymal cells.
Dentin phosphophoryn (DPP) is a major noncollagenous protein in the dentin matrix. In this study, we demonstrate that pluripotent stem cells such as C3H10T1/2 and human bone marrow cells can be committed to the osteogenic lineage by DPP. Treatment with DPP can stimulate the release of intracellular Ca(2+). This calcium flux triggered the activation of Ca(2+)-calmodulin-dependent protein kinase II (CaMKII). Activated CaMKII induced the phosphorylation of Smad1 and promoted nuclear translocation of p-Smad1. Inhibition of store Ca(2+) depletion by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) or down-regulation of CaMKII by KN-62, a selective cell-permeable pharmacological inhibitor or a dominant negative plasmid of CaMKII, blocked DPP-mediated Smad1 phosphorylation. Activation of Smad1 resulted in the expression of osteogenic markers such as Runx2, Osterix, DMP1, Bone sialoprotein, Osteocalcin, NFATc1, and Schnurri-2, which have been implicated in osteoblast differentiation. These findings suggest that DPP is capable of triggering commitment of pluripotent stem cells to the osteogenic lineage.
Department of Oral Biology, University of Illinois, Chicago, Illinois 60612.
Asha
A
Eapen
Department of Oral Biology, University of Illinois, Chicago, Illinois 60612.
Roma
R
Kulkarni
Department of Oral Biology, University of Illinois, Chicago, Illinois 60612.
Sriram
S
Ravindran
Department of Oral Biology, University of Illinois, Chicago, Illinois 60612.
Amsaveni
A
Ramachandran
Department of Pharmacology, University of Illinois, Chicago, Illinois 60612.
Premanand
P
Sundivakkam
Department of Pharmacology, University of Illinois, Chicago, Illinois 60612.
Chinnaswammy
C
Tiruppathi
Department of Oral Biology, University of Illinois, Chicago, Illinois 60612. Electronic address: anneg@uic.edu.
Anne
A
George
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D000943:Antigens, Differentiation / Q000378:metabolism; D020013:Calcium Signaling; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D002417:Cattle; D002454:Cell Differentiation; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004789:Enzyme Activation; D016326:Extracellular Matrix Proteins / Q000502:physiology*; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000201:enzymology* / Q000378:metabolism / Q000502:physiology; D051379:Mice; D050778:NFATC Transcription Factors / Q000235:genetics / Q000378:metabolism; D010006:Osteoblasts / Q000378:metabolism; D010012:Osteogenesis; D010750:Phosphoproteins / Q000502:physiology*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D012795:Sialoglycoproteins / Q000502:physiology*; D051896:Smad Proteins, Receptor-Regulated / Q000378:metabolism; D051898:Smad1 Protein / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D000099311:Dentin Sialophosphoprotein
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000943:Antigens, Differentiation; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D004268:DNA-Binding Proteins; D016326:Extracellular Matrix Proteins; D050778:NFATC Transcription Factors; D010750:Phosphoproteins; D012795:Sialoglycoproteins; D051896:Smad Proteins, Receptor-Regulated; D051898:Smad1 Protein; D014157:Transcription Factors; D000099311:Dentin Sialophosphoprotein; C494003:NFATC1 protein, human; C497399:SMAD1 protein, human; C077171:HIVEP2 protein, human; C063302:KN 62