Address for correspondence: Rocky S. Tuan, Ph.D., Cartilage Biology and Orthopaedics Branch, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, 50 South Drive, Building 50, Room 1503, Bethesda, MD 20892-8022. Voice: 301-451-6854; fax: 301-402-2724.
tuanr{at}mail.nih.gov
Ann. N.Y. Acad. Sci. 961: 172-177 (2002).
Collagenase-treated, explanted human trabecular-bone chips are
an excellent source of osteoblast-like cells. We have recently
shown the multiple differentiation potential of these cells;
in addition to osteogenesis and adipogenesis, these cells also
undergo chondrogenesis when maintained as high-density pellet
cultures (250,000 cells/pellet) in a serum-free, chemically
defined medium stimulated with TGF-ß1 (10 ng/mL).
In this investigation, we have analyzed how transactivating
nuclear transcription factors, specifically AP-2 and SP-1, may
interact with common
cis-acting elements found in the regulatory
region of cartilage-specific genes as part of the signal transduction
mechanism of TGF-ß1 and p38 during chondrogenesis
of human trabecular bone-derived multipotential cells. Both
TGF-ß1 stimulation and p38 MAP kinase activation affect
the binding of AP-2 as well as SP-1 to oligonucleotides with
sequence similarity to the overlapping AP-2/SP-1 sites found
in the putative 52-bp immediate upstream regulatory region and
the 5'-untranslated region of the human aggrecan gene. Electrophoretic
mobility shift assays show that TGF-ß1 treatment of
the bone-derived cells inhibits AP-2 DNA binding but enhances
the DNA binding ability of SP-1. Additionally, treatment of
these TGF-ß1-stimulated cells with p38 MAP kinase
inhibitor, SB203580, rescued the AP-2 DNA binding but did not
affect SP-1 DNA binding. These findings indicate that AP-2 DNA
binding is the target of both TGF-ß1 and p38 MAP kinase
signaling pathways and suggest a possible signal transduction
cascade whereby TGF-ß1 induction of chondrogenesis
involves the activation of p38 MAP kinase and the subsequent
inhibition of DNA binding by AP-2, thereby preventing the transcriptional
repression of the aggrecan gene.
