title: Identification of cis-regulatory sequences in the human angiotensinogen gene by transgene coplacement and site-specific recombination.
abstract: The function of putative regulatory sequences identified in cell transfection experiments can be elucidated only through in vivo experimentation. However, studies of gene regulation in transgenic mice (TgM) are often compromised by the position effects, in which independent transgene insertions differ in expression depending on their location in the genome. In order to overcome such a dilemma, a method called transgene coplacement has been developed in Drosophila melanogaster. In this method, any two sequences can be positioned at exactly the same genomic site by making use of Cre/loxP recombination. Here we applied this method to mouse genetics to characterize the function of direct repeat (DR) sequences in the promoter of the human angiotensinogen (hAGT) gene, the precursor of the vasoactive octapeptide angiotensin II. We modified a hAGT bacterial artificial chromosome to use Cre/loxP recombination in utero to generate TgM lines bearing a wild-type or a mutant promoter-driven hAGT locus integrated at a single chromosomal position. The expression analyses revealed that DR sequences contribute 50 or >95% to hAGT transcription in the liver and kidneys, respectively, whereas same sequences are not required in the heart and brain. This is the first in vivo dissection of DNA cis elements that are demonstrably indispensable for regulating both the level and cell type specificity of hAGT gene transcription.
authors.affiliation: Graduate School of Life and Environmental Sciences, University of Tsukuba, Tennoudai 1-1-1, Tsukuba, Ibaraki 305-8577, Japan. keiji@tara.tsukuba.ac.jp.
authors.forename: Taku
authors.initials: T
authors.lastname: Shimizu
authors.forename: Takayuki
authors.initials: T
authors.lastname: Oishi
authors.forename: Akane
authors.initials: A
authors.lastname: Omori
authors.forename: Akiko
authors.initials: A
authors.lastname: Sugiura
authors.forename: Keiko
authors.initials: K
authors.lastname: Hirota
authors.forename: Hisanori
authors.initials: H
authors.lastname: Aoyama
authors.forename: Tomoko
authors.initials: T
authors.lastname: Saito
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sugaya
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kon
authors.forename: James Douglas
authors.initials: JD
authors.lastname: Engel
authors.forename: Akiyoshi
authors.initials: A
authors.lastname: Fukamizu
authors.forename: Keiji
authors.initials: K
authors.lastname: Tanimoto
mesh_terms: D000808:Angiotensinogen / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D022202:Chromosomes, Artificial, Bacterial; D015536:Down-Regulation; D018390:Gene Targeting / Q000379:methods*; D006801:Humans; D019426:Integrases / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic / Q000235:genetics*; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics*; D014018:Tissue Distribution; D014162:Transfection; D019076:Transgenes; D014764:Viral Proteins / Q000235:genetics
chemical_list: D014764:Viral Proteins; D000808:Angiotensinogen; C045073:Cre recombinase; D019426:Integrases