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This Article Full Text Full Text (PDF) Supplemental Table All Versions of this Article: 295/3/F642    most recent 00384.2007v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Zhou, X. Articles by Sigmund, C. D. PubMed PubMed Citation Articles by Zhou, X. Articles by Sigmund, C. D.

Dysregulated human renin expression in transgenic mice carrying truncated genomic constructs: evidence supporting the presence of insulators at the renin locus

¹Molecular and Cellular Biology Graduate Program, Departments of ²Molecular Physiology and Biophysics and of ³Internal Medicine, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa

Submitted 13 August 2007 ; accepted in final form 9 July 2008

We previously generated transgenic mice carrying a large P1 artificial chromosome (PAC160) encompassing a 160-kb segment containing the human renin gene, two upstream genes, and one downstream gene. We also previously generated mutant PAC160 constructs lacking the distal enhancer and concluded it is required to maintain baseline expression of human renin, but is not required for tissue-specific, cell-specific, and regulated expression of renin in vivo. We now report two additional transgenic lines carrying random truncations of PAC160 upstream of the renin gene. Southern and PCR mapping studies indicate that the truncation break points in the two lines are located 10.4 and 2.5 kb upstream of the renin gene causing a deletion of all DNA upstream of the break. We tested the hypothesis that large-scale deletion of DNA upstream of the human renin gene including the enhancer would cause dysregulation of human renin expression. Phenotypically, these truncations cause a severe dysregulation of human renin expression, but remarkably, a preservation of the normal tissue-specific expression of the human ethanolamine kinase 2 (ETNK2) gene which lies immediately downstream of renin. Several functional binding sites for CTCF, a mammalian insulator protein, were identified in and around the renin and ETNK2 loci by gel shift and chromatin immunoprecipitation. We conclude that there are sequences in and around the renin and ETNK2 loci which act as boundaries between neighboring genes which insulate them from each other. The study illustrates the value of taking a much wider genomic perspective when studying mechanisms regulating gene expression.

P1 artificial chromosome; gene expression; chromatin; transcription