ALU SEQUENCE INVOLVEMENT IN TRANSCRIPTIONAL INSULATION OF THE KERATIN18 GENE IN TRANSGENIC MICE

The human keratin 18 (K18) gene is expressed in a variety of adult sim ple epithelial tissues, including liver, intestine, lung, and kidney, but is not normally found in skin, muscle, heart, spleen, or most of t he brain. Transgenic animals derived from the cloned K18 gene express the transgene in appropriate tissues at levels directly proportional t o the copy number and independently of the sites of integration. We ha ve investigated in transgenic mice the dependence of K18 gene expressi on on the distal 5' and 3' flanking sequences and upon the RNA polymer ase III promoter of an Alu repetitive DNA transcription unit immediate ly upstream of the K18 promoter. Integration site-independent expressi on of tandemly duplicated K18 transgenes requires the presence of eith er an 825-bp fragment of the 5' flanking sequence or the 3.5-kb 3' fla nking sequence. Mutation of the RNA polymerase III promoter of the Alu element within the 825-bp fragment abolishes copy number-dependent ex pression in kidney but does not abolish integration site-independent e xpression when assayed in the absence of the 3' flanking sequence of t he K18 gene. The characteristics of integration site-independent expre ssion and copy number-dependent expression are separable. In addition, the formation of the chromatin state of the K18 gene, which likely re stricts the tissue-specific expression of this gene, is not dependent upon the distal flanking sequences of the 10-kb K18 gene but rather ma y depend on internal regulatory regions of the gene.