The process of chondrogenesis can be mimicked in vitro by insulin treatment of mouse ATDC5 chondroprogenitor cells. To identify novel factors that are involved in the control of chondrogenesis, we carried out a large-scale screening through retroviral insertion mutagenesis and isolated a fast-growing ATDC5 clone incapable of chondrogenic differentiation. Inverse-PCR analysis of this clone revealed that the retroviral DNA was inserted into the promoter region of mouse Id2 (inhibitor of DNA-binding protein 2) gene. This retroviral insertion increased Id2 protein levels to twice those found in normal ATDC5 cells. To investigate whether an elevated level of Id2 protein was responsible for inhibition of chondrogenic differentiation, ATDC5 cells were infected with a retrovirus to stably express Id2. ATDC5 cells expressing ectopic Id2 exhibited signs of de-differentiation, such as rapid growth, and insulin failed to induce expression of Sox9 (Sry-type high-mobility-group box 9) or matrix genes such as type II collagen (COL2) in these cells. When endogenous Id2 was knocked down by siRNA (small interfering RNA) in ATDC5 cells, expression of Sox9 and COL2 was increased and chondrogenic differentiation was accelerated. To examine how Id2 is expressed in chondrocytes in vivo, we carried out immunostaining of E16.5 mouse embryos and found that Id2 is expressed in articular chondrocytes and proliferating chondrocytes, but barely detectable in hypertrophic chondrocytes. Our results suggest that proper expression of Id2 is important to achieving a fine balance between growth and differentiation during chondrogenesis.
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Research Article|
April 14 2009
Proper expression of helix–loop–helix protein Id2 is important to chondrogenic differentiation of ATDC5 cells Available to Purchase
Liu Yang;
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
2To whom correspondence should be addressed: email [email protected]).
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Xiaoyun Ma;
Xiaoyun Ma
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
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Anne Lyone;
Anne Lyone
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
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Junhui Zou;
Junhui Zou
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
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Michael L. Blackburn;
Michael L. Blackburn
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
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Jing Pan;
Jing Pan
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
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Dingqiao Yang;
Dingqiao Yang
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
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Hiroshi Matsushita;
Hiroshi Matsushita
*Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, U.S.A.
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Bin Mei;
Bin Mei
†Department of Orthopedics, University of Washington, 1660 S. Columbian Way, GMR 151, Seattle, WA 98108, U.S.A.
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Anna Zielinska-Kwiatkowska;
Anna Zielinska-Kwiatkowska
†Department of Orthopedics, University of Washington, 1660 S. Columbian Way, GMR 151, Seattle, WA 98108, U.S.A.
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Howard A. Chansky
Howard A. Chansky
†Department of Orthopedics, University of Washington, 1660 S. Columbian Way, GMR 151, Seattle, WA 98108, U.S.A.
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Publisher: Portland Press Ltd
Received:
August 21 2008
Revision Received:
January 16 2009
Accepted:
January 28 2009
Accepted Manuscript online:
January 28 2009
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2009 Biochemical Society
2009
Biochem J (2009) 419 (3): 635–643.
Article history
Received:
August 21 2008
Revision Received:
January 16 2009
Accepted:
January 28 2009
Accepted Manuscript online:
January 28 2009
Citation
Liu Yang, Xiaoyun Ma, Anne Lyone, Junhui Zou, Michael L. Blackburn, Jing Pan, Dingqiao Yang, Hiroshi Matsushita, Bin Mei, Anna Zielinska-Kwiatkowska, Howard A. Chansky; Proper expression of helix–loop–helix protein Id2 is important to chondrogenic differentiation of ATDC5 cells. Biochem J 1 May 2009; 419 (3): 635–643. doi: https://doi.org/10.1042/BJ20081715
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