C/EBPbeta Promotes transition from proliferation to hypertrophic differentiation of chondrocytes through transactivation of p57.
BACKGROUND
Although transition from proliferation to hypertrophic differentiation of chondrocytes is a crucial step for endochondral ossification in physiological skeletal growth and pathological disorders like osteoarthritis, the underlying mechanism remains an enigma. This study investigated the role of the transcription factor CCAAT/enhancer-binding protein beta (C/EBPbeta) in chondrocytes during endochondral ossification.


RESULTS
Mouse embryos with homozygous deficiency in C/EBPbeta (C/EBPbeta-/-) exhibited dwarfism with elongated proliferative zone and delayed chondrocyte hypertrophy in the growth plate cartilage. In the cultures of primary C/EBPbeta-/- chondrocytes, cell proliferation was enhanced while hypertrophic differentiation was suppressed. Contrarily, retroviral overexpression of C/EBPbeta in chondrocytes suppressed the proliferation and enhanced the hypertrophy, suggesting the cell cycle arrest by C/EBPbeta. In fact, a DNA cell cycle histogram revealed that the C/EBPbeta overexpression caused accumulation of cells in the G0/G1 fraction. Among cell cycle factors, microarray and real-time RT-PCR analyses have identified the cyclin-dependent kinase inhibitor p57(Kip2) as the transcriptional target of C/EBPbeta. p57(Kip2) was co-localized with C/EBPbeta in late proliferative and pre-hypertrophic chondrocytes of the mouse growth plate, which was decreased by the C/EBPbeta deficiency. Luciferase-reporter and electrophoretic mobility shift assays identified the core responsive element of C/EBPbeta in the p57(Kip2) promoter between -150 and -130 bp region containing a putative C/EBP motif. The knockdown of p57(Kip2) by the siRNA inhibited the C/EBPbeta-induced chondrocyte hypertrophy. Finally, when we created the experimental osteoarthritis model by inducing instability in the knee joints of adult mice of wild-type and C/EBPbeta+/- littermates, the C/EBPbeta insufficiency caused resistance to joint cartilage destruction.


CONCLUSIONS
C/EBPbeta transactivates p57(Kip2) to promote transition from proliferation to hypertrophic differentiation of chondrocytes during endochondral ossification, suggesting that the C/EBPbeta-p57(Kip2) signal would be a therapeutic target of skeletal disorders like growth retardation and osteoarthritis.
Hirata
Makoto
M
Department of Sensory & Motor System Medicine, Faculty of Medicine, University of Tokyo, Tokyo, Japan.
Kugimiya
Fumitaka
F
Fukai
Atsushi
A
Ohba
Shinsuke
S
Kawamura
Naohiro
N
Ogasawara
Toru
T
Kawasaki
Yosuke
Y
Saito
Taku
T
Yano
Fumiko
F
Ikeda
Toshiyuki
T
Nakamura
Kozo
K
Chung
Ung-Il
UI
Kawaguchi
Hiroshi
H
D000818:Animals; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000502:physiology*; D002453:Cell Cycle / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics*; D049109:Cell Proliferation; D019902:Chondrocytes / Q000166:cytology*; D050761:Cyclin-Dependent Kinase Inhibitor p57 / Q000235:genetics* / Q000502:physiology; D004622:Embryo, Mammalian; D051379:Mice; D010003:Osteoarthritis / Q000235:genetics; D010012:Osteogenesis; D034741:RNA, Small Interfering / Q000494:pharmacology; D015533:Transcriptional Activation
D022782:CCAAT-Enhancer-Binding Protein-beta; C493965:Cdkn1c protein, mouse; D050761:Cyclin-Dependent Kinase Inhibitor p57; D034741:RNA, Small Interfering