Epithelial Systems Biology Laboratory, Systems Biology Center, National Heart, Lung, and Blood Institute , Bethesda, Maryland.
Department of Regenerative and Cancer Cell Biology, Albany Medical College , Albany, New York.
Am J Physiol Renal Physiol. 2018 Apr 1;314(4):F572-F583. doi: 10.1152/ajprenal.00436.2017. Epub 2017 Dec 20.
An ideal inducible system should be cell specific and have absolutely no background recombination without induction (i.e., no leakiness), a high recombination rate after induction, and complete fidelity in cell specificity (i.e., restricted recombination exclusively in cells where the driver gene is expressed). However, such an ideal mouse model remains unavailable for collecting duct research. Here, we report a mouse model that meets these criteria. In this model, a cassette expressing ERCreER ( ECE) is inserted at the ATG of the endogenous Aqp2 locus to disrupt Aqp2 function and to express ECE under the control of the Aqp2 promoter. The resulting allele is named Aqp2. There was no indication of a significant impact of disruption of a copy of Aqp2 on renal function and blood pressure control in adult Aqp2 heterozygotes. Without tamoxifen, Aqp2 did not activate a Cre-dependent red fluorescence protein (RFP) reporter in adult kidneys. A single injection of tamoxifen (2 mg) to adult mice enabled Aqp2 to induce robust RFP expression in the whole kidney 24 h postinjection, with the highest recombination efficiency of 95% in the inner medulla. All RFP-labeled cells expressed principal cell markers (Aqp2 and Aqp3), but not intercalated cell markers (V-ATPase B1B2, and carbonic anhydrase II). Hence, Aqp2 confers principal cell-specific tamoxifen-inducible recombination with absolutely no leakiness, high inducibility, and complete fidelity in cell specificity, which should be an important tool for temporospatial control of target genes in the principal cells and for Aqp2 lineage tracing in adult mice.
理想的诱导系统应该具有细胞特异性，在没有诱导的情况下绝对没有背景重组（即无渗漏），诱导后重组率高，并且在细胞特异性上具有完全的保真度（即驱动基因表达的细胞中仅发生限制性重组）。然而，这种理想的小鼠模型仍然无法用于收集管研究。在这里，我们报告了一种符合这些标准的小鼠模型。在该模型中，一个表达 ERCreER（ECE）的盒在内源性 Aqp2 基因座的 ATG 处插入，以破坏 Aqp2 的功能，并在 Aqp2 启动子的控制下表达 ECE。由此产生的等位基因命名为 Aqp2。在成年 Aqp2 杂合子中，破坏 Aqp2 一个拷贝对肾功能和血压控制没有明显影响的迹象。没有他莫昔芬，Aqp2 不会在成年肾脏中激活 Cre 依赖性红色荧光蛋白（RFP）报告基因。单次给予成年小鼠 2mg 他莫昔芬，可使 Aqp2 在注射后 24 小时内在整个肾脏中诱导强烈的 RFP 表达，在内髓中的重组效率最高达 95%。所有 RFP 标记的细胞均表达主细胞标记物（Aqp2 和 Aqp3），但不表达闰细胞标记物（V-ATPase B1B2 和碳酸酐酶 II）。因此，Aqp2 赋予主细胞特异性他莫昔芬诱导重组，绝对无渗漏，诱导率高，细胞特异性完全保真，这应该是在主细胞中时空控制靶基因和 Aqp2 谱系追踪的重要工具成年小鼠。
