title: Phosphophoryn regulates the gene expression and differentiation of NIH3T3, MC3T3-E1, and human mesenchymal stem cells via the integrin/MAPK signaling pathway.
abstract: Extracellular matrix proteins (ECMs) serve as both a structural support for cells and a dynamic biochemical network that directs cellular activities. ECM proteins such as those of the SIBLING family (small integrin-binding ligand glycoprotein) could possess inherent growth factor activity. In this study, we demonstrate that exon 5 of dentin matrix protein 3 (phosphophoryn (PP)), a non-collagenous dentin ECM protein and SIBLING protein family member, up-regulates osteoblast marker genes in primary human adult mesenchymal stem cells (hMSCs), a mouse osteoblastic cell line (MC3T3-E1), and a mouse fibroblastic cell line (NIH3T3). Quantitative real-time PCR technology was used to quantify gene expression levels of bone markers such as Runx2, Osx (Osterix), bone/liver/kidney Alp (alkaline phosphatase), Ocn (osteocalcin), and Bsp (bone sialoprotein) in response to recombinant PP and stably transfected PP. PP up-regulated Runx2, Osx, and Ocn gene expression. PP increased OCN protein production in hMSCs and MC3T3-E1. ALP activity and calcium deposition was increased by PP in hMSC. Furthermore, an alpha(v)beta(3) integrin-blocking antibody significantly inhibited recombinant PP-induced expression of Runx2 in hMSCs, suggesting that signaling by PP is mediated through the integrin pathway. PP was also shown to activate p38, ERK1/2, and JNK, three components of the MAPK pathway. These data demonstrate a novel signaling function for PP in cell differentiation beyond the hypothesized role of PP in biomineralization.
authors.affiliation: Department of Oral Medicine and Pathology, School of Dental Medicine, University of Pittsburgh, 3501 Terrace St., Pittsburgh, PA 15261-1964, USA.
authors.forename: Julie
authors.initials: J
authors.lastname: Jadlowiec
authors.forename: Hannjörg
authors.initials: H
authors.lastname: Koch
authors.forename: Xiaoyuan
authors.initials: X
authors.lastname: Zhang
authors.forename: Phil G
authors.initials: PG
authors.lastname: Campbell
authors.forename: Merriam
authors.initials: M
authors.lastname: Seyedain
authors.forename: Charles
authors.initials: C
authors.lastname: Sfeir
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D050659:Core Binding Factor Alpha 1 Subunit; D017931:DNA Primers / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000378:metabolism; D003804:Dentin / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005091:Exons; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D058574:Integrin-Binding Sialoprotein; D016023:Integrins / Q000378:metabolism; D020935:MAP Kinase Signaling System; D008648:Mesoderm / Q000166:cytology; D051379:Mice; D008954:Models, Biological; D041681:NIH 3T3 Cells; D015675:Osteocalcin / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012795:Sialoglycoproteins / Q000378:metabolism; D015398:Signal Transduction; D000076182:Sp7 Transcription Factor; D013234:Stem Cells; D013997:Time Factors; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000378:metabolism; D015854:Up-Regulation
chemical_list: D050659:Core Binding Factor Alpha 1 Subunit; D017931:DNA Primers; D004268:DNA-Binding Proteins; C546871:IBSP protein, human; C546872:Ibsp protein, mouse; D058574:Integrin-Binding Sialoprotein; D016023:Integrins; D010750:Phosphoproteins; D011506:Proteins; C493709:RUNX2 protein, human; D011994:Recombinant Proteins; C493716:Runx2 protein, mouse; D012795:Sialoglycoproteins; D000076182:Sp7 Transcription Factor; C478163:SP7 protein, human; C479466:Sp7 protein, mouse; D050656:Transcription Factor AP-2; D014157:Transcription Factors; C017630:phosphophoryn; D015675:Osteocalcin; D000469:Alkaline Phosphatase; D002118:Calcium