Regulatory mechanism of human connective tissue growth factor (CTGF/Hcs24)gene expression in a human chondrocytic cell line, HCS-2/8

CTGF/Hcs24 is a multi-functional growth factor that potentiates either the growth or differentiation of mesenchymal cells, according to the biological conditions. Among various functional aspects of CTGF/Hcs24, it is especial ly notable that CTGF/Hcs24 may promote endochondral ossification in growth cartilage through all stages, and it is highly expressed in a human chondro sarcoma-derived chondrocytic cell line (HCS-2/8). In this study, to clarify the regulatory mechanism of CTGF/Hcs24 gene expression in chondrocytes, we analyzed the transcriptional activity of the CTGF/Hcs24 promoter and the e ffect of the CTGF/Hcs24 S'-untranslated region (3'-UTR) on gene expression in HCS-2/8 by means of an established DNA transfection and luciferase repor ter gene assay system. As a result, the luciferase activity of the CTGF/Hcs 24 promoter was found to be remarkably high in HCS-2/8. The 3'-UTR of the C TGF/Hcs24 gene strongly repressed the luciferase activity in HCS-2/8, when it was linked to the downstream of the luciferase reporter gene, suggesting its functionality also in chondrocytic cells. Deletion analysis of the CTG F/Hcs24 promoter clarified a major segment responsible for the enhanced CTG F/Hcs24 promoter activity in HCS-2/8, The TGF-beta response element in the DNA segment was active in HCS-2/8, and point mutations in the element moder ately decreased the highly maintained promoter activity with total loss of TGF-beta responsiveness. These results indicate that the strong expression of the CTGF/Hcs24 gene in HCS-2/8 was mainly caused by high transcriptional activity of the CTGF/Hcs24 promoter, and that the TGF-beta response elemen t is one of the critical elements that support the high transcription activ ity.