Genomic organization of the human chondromodulin-1 gene containing a promoter region that confers the expression of reporter gene in chondrogenic ATDC5 cells

Chondromodulin-1 (ChM-1) is a cartilage-specific glycoprotein that stimulat es the growth of chondrocytes and inhibits the tube formation of endothelia l cells. To clarify the tissue-specific expression and the role of ChM-1 in pathophysiological conditions, we analyzed the structure of the human ChM- 1 gene and its promoter. On the screening of a human genomic cosmid library using the human ChM-1 complimentary DNA (cDNA) as a probe, two clones were obtained that contained ChM-1 cDNA, The restriction enzyme map and nucleot ide sequence revealed the human ChM-1 gene consisting: of seven exons and e xon-intron boundaries. The human ChM-1 gene was assigned to chromosome 13q1 4-21 by fluorescence in situ hybridization (FISH) using the clone as a prob e. A primer extension analysis using total RNA extracted from human cartila ge revealed a major transcription start site with the sequence CGCT(+1)GG. The region approximately 3-kilobase (kb) nucleotides upstream of the transl ation start site was then sequenced and analyzed in terms of promoter activ ity. We found that a region 446 base pairs (bp) upstream of the start site had promoter activity in COS7, HeLa, and ATDC5 cells. In structure the prom oter is a TATA-less type without a CC-rich region.-The transcription factor s Sox9, Og12, and Cart-1 did not affect the promoter activity,The transcrip tion factor Ying-Yang1 suppressed the promoter activity but GABP protein di d not change the promoter activity. The construct containing -446/+87 fused to the SV40 enhancer and green fluorescent protein (CFP) exhibited express ion of CFP corresponding to the differentiation of ATDC5 cells to mature ch ondrocytes, These results suggest that the element -446/+87 confers the car tilage-specific expression of this gene by some factor(s) other than Sox9, Og12, and Cart-1.