Pitx2 prevents osteoblastic transdifferentiation of myoblasts by bone morphogenetic proteins.
Muscle cells are often exposed to bone morphogenetic proteins (BMPs) in pathological muscle and/or bone conditions. Because BMPs function as strong bone inducers as well as myogenesis inhibitors, certain molecules likely prevent muscle cells from converting into pathologic bone; without these molecules, de novo bone would form as observed in myositis ossificans traumatica. When C2C12 myoblasts are exposed to BMPs, they differentiate into osteoblastic cells but cannot mature into bone cells. As the Osterix gene, a transcription factor for osteoblast differentiation, is only transiently induced upon BMP stimulation in C2C12 cells, we hypothesized that unknown transcriptional repressor(s) inhibit Osterix expression and prevent complete osteoblastic differentiation. Gene microarray analyses were performed to identify putative inhibitors for osteoblastic differentiation, and the paired-like homeodomain transcription factor Pitx2 (also termed Rieg), which plays an important regulatory role in left-right asymmetry, was identified. Pitx2 was induced 2 days after BMP stimulation in C2C12 cells in concert with Osterix down-regulation. Overexpression of Pitx2 repressed Osterix expression and subsequent osteoblastic differentiation, whereas Runx2, the most upstream regulator of osteogenesis, was unaffected. Conversely, the induction of short hairpin RNA for Pitx2 in C2C12 cells enhanced Osterix expression and osteoblastic maturation upon BMP stimulation. Moreover, mouse embryonic fibroblasts containing myoblasts from Pitx2-null embryos showed enhanced Osterix expression upon BMP stimulation. These findings suggest that Pitx2 suppresses osteogenic signals induced by BMPs in myoblasts to prevent their osteoblastic conversion.
Hayashi
Makoto
M
Department of Biochemistry, The Cancer Institute of the Japanese Foundation for Cancer Research, 3-10-6 Ariake, Koto-ku, Tokyo 135-8550, Japan; Department of Biological Sciences, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan.
Maeda
Shingo
S
Department of Biochemistry, The Cancer Institute of the Japanese Foundation for Cancer Research, 3-10-6 Ariake, Koto-ku, Tokyo 135-8550, Japan. Electronic address: shingo.maeda@jfcr.or.jp.
Aburatani
Hiroyuki
H
Genome Science Division, RCAST, University of Tokyo, 4-6-1 Komaba, Meguro-ku, Tokyo 153-8904, Japan.
Kitamura
Kunio
K
Department of Mental Retardation and Birth Defect Research, National Institute of Neuroscience, National Center of Neurology and Psychiatry, 4-1-1 Ogawahigashi, Kodaira, Tokyo 187-8502, Japan.
Miyoshi
Hiroyuki
H
Subteam for Manipulation of Cell Fate, BioResource Center, RIKEN, 3-1-1 Koyadai, Tsukuba-shi, Ibaraki 305-0074, Japan.
Miyazono
Kohei
K
Department of Molecular Pathology, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Imamura
Takeshi
T
Department of Biochemistry, The Cancer Institute of the Japanese Foundation for Cancer Research, 3-10-6 Ariake, Koto-ku, Tokyo 135-8550, Japan.
D000818:Animals; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology*; D002460:Cell Line; D054338:Cell Transdifferentiation / Q000187:drug effects*; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D032446:Myoblasts / Q000166:cytology / Q000187:drug effects*; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects*; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D000076182:Sp7 Transcription Factor; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D000097577:Homeobox Protein PITX2
D019485:Bone Morphogenetic Proteins; D018398:Homeodomain Proteins; D034741:RNA, Small Interfering; D000076182:Sp7 Transcription Factor; D014157:Transcription Factors; D000097577:Homeobox Protein PITX2; C479466:Sp7 protein, mouse