HUMAN CART-1 - STRUCTURAL ORGANIZATION, CHROMOSOMAL LOCALIZATION, ANDFUNCTIONAL-ANALYSIS OF A CARTILAGE-SPECIFIC HOMEODOMAIN CDNA

Homeoproteins control cell fates during development, specifying patter n formation and the ontogeny of specific tissues and organs in embryog enesis. Cart-1 cDNA was recently cloned from a rat chondrosarcoma tumo r and it encodes a protein containing a paired-like homeodomain that i s selectively expressed in cartilage during early chondrocyte differen tiation. Here we report the molecular cloning of the human Cart-1 cDNA from a HeLa cervical carcinoma cDNA library. The human Cart-1 cDNA se quence is 88% identical and the deduced amino acid sequence is 95% ide ntical to the rat sequence, indicating that Cart-1 structure is highly conserved, Northern and reverse transcriptase polymerase chain reacti on (RT-PCR) analysis revealed Cart-1 mRNA expression in HeLa cervical carcinoma cells and human cervical tissue, but Cart-1 mRNA was not det ected in GH(3) rat pituitary cells and murine 10T1/2 one-half fibrobla st cells, The Cart-1 gene was localized to human chromosome 12 and reg ionally mapped to the 12q21.3-q22 by PCR analysis of rodent-x-human so matic cell hybrid DNA and the CEPH megabase-insert YAC DNA pools, resp ectively, The Holt-Gram syndrome, characterized by upper limb and atri al septal dysplasias, also maps to the 12q21.3-q22 region. Cotransfect ion studies show that Cart-1 inhibits the rat prolactin promoter and t hat this repression is mediated by footprint II, an AT-rich element th at functions as an inhibitory site of prolactin gene expression in non pituitary cells and which was used to clone Cart-1. Taken together, th ese data indicate that Cart-1 may also influence cervix development, i dentify a putative DNA binding site for Cart-1, and, begin to define i ts functional role as modulator of gene expression.