A Kr  ppel-associated box- XXXX finger protein, @entity70895 , represses cell-type-specific promoter activity of the alpha 2(XI) collagen gene.

@entity70895 @entity1 @entity375 @entity511 @entity19 @entity1184 @entity70896 @entity219 @entity43263 @entity39529
Type XI collagen is composed of three chains, alpha 1(XI), alpha 2(XI), and alpha 3(XI), and plays a critical role in the formation of @entity39529 fibrils and in skeletal morphogenesis. It was previously reported that the -530-bp promoter segment of the alpha 2(XI) collagen gene ( @entity43263 ) was sufficient for cartilage-specific expression and that a 24-bp sequence from this segment was able to switch promoter activity from neural tissues to cartilage in @entity19 when this sequence was placed in the heterologous neurofilament light gene (NFL) promoter. To identify a protein factor that bound to the 24-bp sequence of the @entity43263 promoter, we screened a @entity19 limb bud cDNA expression library in the @entity375 one-hybrid screening system and obtained the cDNA clone @entity70895 . Sequence analysis revealed that @entity70895 is a @entity219 finger protein consisting of a Kr  ppel-associated box (KRAB) and is a homologue of @entity1 @entity70896 , which was previously isolated by random cloning and sequencing. The KRAB domain has been found in a number of @entity219 finger proteins and implicated as a transcriptional repression domain, although few target genes for KRAB-containing @entity219 finger proteins has been identified. Here, we demonstrate that @entity70895 functions as a negative regulator of @entity43263 . In situ hybridization analysis of developing @entity19 cartilage showed that @entity70895 mRNA is highly expressed by @entity1184 chondrocytes but is minimally expressed by resting and proliferating chondrocytes, in an inverse correlation with the expression patterns of @entity43263 . Gel shift assays showed that @entity70895 bound a specific sequence within the 24-bp site of the @entity43263 promoter. We found that @entity43263 promoter activity was inhibited by transfection of the @entity70895 expression vector in RSC cells, a @entity511 cell line. The expression vector for mutant @entity70895 lacking the KRAB domain failed to inhibit @entity43263 promoter activity. These results demonstrate that KRAB- @entity219 finger protein @entity70895 inhibits transcription of its physiological target gene, suggesting a novel regulatory mechanism of cartilage-specific expression of @entity43263 .
