Identification of the cyclin D1 gene as a target of activating transcription factor 2 in chondrocytes

Endochondral bone growth is regulated by the rates of chondrocyte prolifera tion and differentiation. However, the intracellular mechanisms regulating these processes are poorly understood. Recently, interruption of the gene e ncoding the transcription factor activating transcription factor 2 (ATF-2) was shown to inhibit proliferation of chondrocytes in mice [Reimold, A. M,, et al, (1996) Nature (London) 379, 262-265], The target genes of ATF-2 tha t are responsible for this phenotype remain unknown, Here we report that th e cyclin D1 gene is a direct target of ATF-2 in chondrocytes, ATF-2 is pres ent in nuclear extracts from chondrogenic cell lines and binds, as a comple x with a CRE-binding protein (CREB)/CRE modulator protein, to the cAMP resp onse element (CRE) in the cyclin D1 promoter. Mutation of the cyclin D1 CRE caused a 78% reduction in the activity of the promoter in chondrocytes. Ov erexpression of ATF-2 in chondrocytes enhanced activity of the cyclin D1 pr omoter 3.5-fold. In contrast, inhibition of endogenous ATF-2 or CREB by exp ression of dominant-negative inhibitors of CREB and ATF-2 significantly red uced the activity of the promoter in chondrocytes through the CRE, In addit ion, levels of cyclin D1 protein are greatly reduced in the chondrocytes of ATF-2-deficient mice. These data identify the cyclin D1 gene as a direct t arget of ATF-2 in chondrocytes and suggest that reduced expression of cycli n D1 contributes to the defective cartilage development of these mice.