Cre-mediated modulation of gene function in the murine retinal pigment epithelium (RPE) has been widely used, but current postnatal RPE-selective Cre driver lines suffer from limited recombination efficiency and/or ectopic or mosaic expression. We sought to generate a transgenic mouse line with consistently efficient RPE-selective Cre activity that could be temporally regulated. We used φC31 integrase to insert a DNA construct encoding a human BEST1 promoter fragment driving a Cre recombinase estrogen receptor fusion (BEST1-CreERT2) at the Rosa26 locus of C57BL/6J mice. Rosa26^{BEST1-CreERT2} mice were bred with a tdTomato reporter line and to mice with a Cre-conditional allele of Tfam. 4-hydroxytamoxifen or vehicle was delivered by four consecutive daily intraperitoneal injections. TdTomato was robustly expressed in the RPE of mice of both sexes for inductions beginning at P14 (males 90.7 ± 4.5%, females 84.7 ± 3.2%) and at 7 weeks (males 84.3 ± 7.0%, females 82 ± 3.6%). <0.6% of Muller glia also expressed tdTomato, but no tdTomato fluorescence was observed in other ocular cells or in multiple non-ocular tissues, with the exception of sparse foci in the testis. No evidence of retinal toxicity was observed in mice homozygous for the transgene induced beginning at P14 and assessed at 7–10 months. RPE-selective ablation of Tfam beginning at P14 led to reduced retinal thickness at 8 months of age and diminished retinal electrical responses at 12 months, as expected. These findings demonstrate that we have generated a mouse line with consistently efficient, tamoxifen-mediated postnatal induction of Cre recombination in the RPE and a small fraction of Muller glia. This line should be useful for temporally regulated modulation of gene function in the murine RPE.

Cre 介导的小鼠视网膜色素上皮 (RPE) 中基因功能的调节已被广泛使用，但目前的出生后 RPE 选择性 Cre 驱动线的重组效率和/或异位或镶嵌表达有限。我们试图生成具有持续有效的 RPE 选择性 Cre 活性的转基因小鼠品系，该活性可以在时间上进行调节。我们使用 φC31 整合酶在C57BL/6J 小鼠的Rosa26基因座插入编码人类BEST1启动子片段的 DNA 构建体，该片段驱动 Cre 重组酶雌激素受体融合 ( BEST1-CreERT2 ) 。Rosa26 ^{BEST1-CreERT2}小鼠用 tdTomato 报告基因系和带有Tfam的 Cre 条件等位基因的小鼠繁殖. 4-羟基三苯氧胺或载体通过连续四次每日腹膜内注射递送。TdTomato 在两性小鼠的 RPE 中强烈表达，用于从 P14（雄性 90.7 ± 4.5%，雌性 84.7 ± 3.2%）和第 7 周（雄性 84.3 ± 7.0%，雌性 82 ± 3.6%）开始的诱导。<0.6% 的穆勒神经胶质也表达 tdTomato，但在其他眼细胞或多个非眼组织中未观察到 tdTomato 荧光，但睾丸中的稀疏病灶除外。在 P14 开始诱导并在 7-10 个月评估的转基因纯合小鼠中没有观察到视网膜毒性的证据。正如预期的那样，从 P14 开始对 Tfam 进行 RPE 选择性消融导致 8 个月大时视网膜厚度减少，12 个月时视网膜电反应减弱。这些发现表明，我们已经生成了一种小鼠品系，该品系具有持续有效的、他莫昔芬介导的出生后诱导 RPE 和一小部分穆勒神经胶质中的 Cre 重组。这条线应该可用于小鼠 RPE 中基因功能的时间调节调制。