DIFFERENTIAL REGULATION OF COL2A1 EXPRESSION IN DEVELOPING AND MATURECHONDROCYTES

To investigate the regulation of type II collagen gene expression in c ells undergoing chondrogenic differentiation, we have employed a 5-kbp genomic fragment of the human type II collagen gene which contains 1. 8 kbp of upstream sequences, the transcription start site, the first e xon and 3 kbp of intronic sequences, fused to either lac Z or chloramp henicol acetyl transferase-reporter gene. Transient expression studies revealed a parallel increase in transgene activity and endogenous typ e II collagen mRNA levels during the onset of the cartilage differenti ation of limb mesenchymal cells in high-density micromass cultures. At later periods in culture, however, the transgene activity declines, a lthough steady-state levels of type II collagen mRNA are reported to c ontinue to increase (Kosher et al.: J. Cell. Biol. 202: 1151-1156, 198 6; Kravis and Upholt. Dev. Biol. 108: 164-172, 1985). In addition, the activity of the transgene is seven-fold higher at the onset of chondr ogenic differentiation in micromass cultures than in well differentiat ed sternal chondrocytes, although similar levels of type II collagen t ranscripts are found in these cells. Furthermore, deletions of introni c segments resulted in greater drop in activity of the constructs in d ifferentiating chondrocytes in micromass cultures than in mature stern al chondrocytes. The expression of the construct in transgenic mice is higher at the onset of chondrogenic differentiation and in newly diff erentiated chondrocytes than in more mature differentiated chondrocyte s. Based on these observations, it appears that the mechanisms involve d in the regulation of the type II collagen gene at the onset of chond rocyte differentiation are different from those resulting in the maint ainance of its expression in fully differentiated chondrocytes.