Transcriptional regulation of osteopontin gene in vivo by PEBP2alphaA/CBFA1 and ETS1 in the skeletal tissues.
Osteopontin (Opn) and polyoma enhancer-binding protein (PEBP) 2alphaA/core binding factor (CBFA) 1 have been suggested to play important roles in ossification. The overlapping localization of opn and PEBP2alphaA/CBFA1 mRNA, and the marked decrease of opn mRNA expression in PEBP2alphaA knockout mice, indicated that the transcription of opn gene was controlled by PEBP2alphaA. In the present study, we determined the direct regulation of PEBP2alphaA on the opn promoter activity. Opn promoter activity was markedly enhanced by PEBP2alphaA and ETS1 in a synergistic manner. The synergistic effect was diminished when either the PEBP2alphaA or ETS1 binding site was mutated, or the spatial arrangement of these sites was mutated by a 4-nt insertion. The distance between these sites was important for transactivation but not protein-DNA binding. The direct interaction between PEBP2alphaA and ETS1 was depended on protein-DNA binding. These results suggested that the specific spatial arrangement of both sites and direct interaction between PEBP2alphaA and ETS1, were essential for promoter function. Furthermore, endogenous opn mRNA was decreased with the introduction of dominant negative PEBP2alphaA to MC3T3/E1 cells expressing endogenous PEBP2alphaA, ETS1 and opn. These findings suggest that PEBP2alphaA and ETS1 cooperate in vivo to regulate expression of the opn gene in the skeletal tissue. Cell type-specific regulation of Opn gene expression will also be discussed.
Sato
M
M
Department of Pathology, Osaka University Medical School, Suita, Japan.
Morii
E
E
Komori
T
T
Kawahata
H
H
Sugimoto
M
M
Terai
K
K
Shimizu
H
H
Yasui
T
T
Ogihara
H
H
Yasui
N
N
Ochi
T
T
Kitamura
Y
Y
Ito
Y
Y
Nomura
S
S
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D050659:Core Binding Factor Alpha 1 Subunit; D004268:DNA-Binding Proteins / Q000502:physiology*; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D010012:Osteogenesis / Q000235:genetics; D053495:Osteopontin; D011401:Promoter Regions, Genetic; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D050783:Proto-Oncogene Proteins c-ets; D016415:Sequence Alignment; D012795:Sialoglycoproteins / Q000235:genetics* / Q000378:metabolism; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D050659:Core Binding Factor Alpha 1 Subunit; D004268:DNA-Binding Proteins; C494041:ETS1 protein, human; C494042:Ets1 protein, mouse; D009363:Neoplasm Proteins; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; C493709:RUNX2 protein, human; C493716:Runx2 protein, mouse; C506895:SPP1 protein, human; D012795:Sialoglycoproteins; C506896:Spp1 protein, mouse; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D053495:Osteopontin