A single nucleotide mutation in the mouse renin promoter disrupts blood pressure regulation
Keiji Tanimoto, … , Kazuyuki Yanai, Akiyoshi Fukamizu
Keiji Tanimoto, … , Kazuyuki Yanai, Akiyoshi Fukamizu
Published February 7, 2008
Citation Information: J Clin Invest. 2008;118(3):1006-1016. https://doi.org/10.1172/JCI33824.
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Research Article Nephrology

A single nucleotide mutation in the mouse renin promoter disrupts blood pressure regulation

Abstract

Renin, a major regulatory component of the renin-angiotensin system, plays a pivotal role in regulating blood pressure and electrolyte homeostasis and is predominantly expressed in the kidney. Several cAMP-responsive elements have been identified within renin gene promoters. Here, we study how 2 such elements, renin proximal promoter element-2 (RP-2) and overlapping cAMP and negative regulatory elements (CNRE), affect the transcriptional regulation of renin. We generated Tg mice (TgM) bearing BACs containing either WT or mutant RP-2 or CNRE, integrated at single chromosomal loci. Analysis of the TgM revealed that RP-2 was essential to basal promoter activity in the kidney, while renin mRNA levels did not significantly change in any tissues tested in the CNRE mutant TgM. To evaluate the physiological significance of these mutations, we used the BAC Tg to rescue hypotensive Renin-null mutant mice. As predicted, no renin expression was observed in the kidneys of RP-2 mutant/Renin-null compound mice, whereas renin expression in CNRE mutant compound mice was indistinguishable from that in control mice. Consistent with this, RP-2 mutant animals were hypotensive, while CNRE mutants had normal blood pressure. Thus, transcriptional regulation of renin expression via RP-2 but not CNRE is critical for blood pressure regulation by this gene.

Authors

Keiji Tanimoto, Akiko Sugiura, Sumiyo Kanafusa, Tomoko Saito, Naoto Masui, Kazuyuki Yanai, Akiyoshi Fukamizu

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Figure 2

Structural analysis of the Tg lines.

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Structural analysis of the Tg lines. (A) Comparison of the Tg Ren-1C and...
(A) Comparison of the Tg Ren-1C and endogenous Ren-2 and Ren-1D loci. The renin genes and SfiI sites are shown as filled boxes and vertical lines, respectively. The artificially introduced SfiI site is marked by a lollipop. Expected fragment sizes (in kbp) after complete enzyme digestion are shown above the line. The probes used for Southern blot analysis in B and C are indicated by solid rectangles (II–VI). (B and C) Southern blot analyses of the TgM carrying either CNRE (B) or RP-2 (C) modifications. Thymic cells from WT (W) and mut (M) TgM lines were embedded in agarose plugs, digested with SfiI, and separated by PFG electrophoresis. DNA blots were hybridized separately to probes (II–VI) shown in A. The sizes of the expected bands are indicated (in kbp) on the left of each panel. Those expected from the endogenous Ren2 and Ren-1D loci are in parentheses, and partially digested DNA is marked by asterisks. (D and E) Fine structural analyses of the CNRE (D) and RP-2 (E) promoter regions. (Left panels) In vivo Cre/loxP recombination removes the Kanr gene plus either mut or WT promoter sequences from the parental locus to generate either the WT or the mut locus, respectively. E, EcoRI; N, NcoI. (Right panels) Tail DNA from parental, WT, and mut TgM was digested with EcoRI (top) or NcoI (bottom), separated on agarose gels, transferred to a nylon membrane, and hybridized with probes 1 and 2 (shaded rectangles in the left panels), respectively.