Two of four alternatively spliced isoforms of RUNX2 control osteocalcin gene expression in human osteoblast cells.
Runx2 is a Runt domain transcription factor that transcriptionally regulates osteoblast differentiation and bone formation. In this study, we show that human chondro- and osteosarcoma cell lines, human mesenchymal stem cells (hMSC) and a human primary chondrocytes (HC), osteoblst cells (HOb) express an intact isoform (RUNX2wt) and 3 alternatively spliced isoforms (RUNX2Delta5, Delta7, and Delta5Delta7) that are generated by skipping exon 5 and/or exon 7. Two of the truncated forms of RUNX2 (RUNX2Delta5 and RUNX2Delta5Delta7) did not localize in the nucleus and had lost their DNA binding activity. In cotransfection experiments with an osteocalcin (OC) promoter construct, we confirmed that only RUNX2wt and RUNX2Delta7 could upregulate the OC promoter activity in the osteosarcoma cell line. In addition, the coactivator CBP/p300 enhanced the transcriptional activity of the OC promoter when coexpressed with RUNX2wt or RUNX2Delta7, but not when coexpressed with RUNX2Delta5 or RUNX2Delta5Delta7. In contrast, the corepressor HDAC3 only repressed the activation from the OC promoter when coexpressed with RUNX2wt. These results support the hypothesis that RUNX2 both up- and downregulates its target gene promoters, as exemplified by the OC gene, using various isoforms and context-dependent formation of transcriptional complexes.
Makita
Naoyuki
N
Research Institute for Biological Sciences, Tokyo University of Science, Chiba, Japan.
Suzuki
Mitsuhiro
M
Asami
Shiori
S
Takahata
Rintaroh
R
Kohzaki
Daika
D
Kobayashi
Sho
S
Hakamazuka
Takashi
T
Hozumi
Nobumichi
N
D017398:Alternative Splicing; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism; D002813:Chondrosarcoma / Q000235:genetics / Q000378:metabolism; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics* / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D005786:Gene Expression Regulation; D006655:Histone Deacetylases / Q000378:metabolism; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D015675:Osteocalcin / Q000235:genetics*; D012516:Osteosarcoma / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D050880:p300-CBP Transcription Factors / Q000378:metabolism; D000099186:Histone Deacetylase 3
D050659:Core Binding Factor Alpha 1 Subunit; D017931:DNA Primers; D006655:Histone Deacetylases; D015675:Osteocalcin; D020033:Protein Isoforms; D011994:Recombinant Proteins; D050880:p300-CBP Transcription Factors; D000099186:Histone Deacetylase 3; C493709:RUNX2 protein, human