Chu Xuran, Taghizadeh Sara, Vazquez-Armendariz Ana Ivonne, Herold Susanne, Chong Lei, Chen Chengshui, Zhang Jin-San, El Agha Elie, Bellusci Saverio
School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, China.
Key Laboratory of Interventional Pulmonology of Zhejiang Province, Department of Pulmonary and Critical Care Medicine, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.
Front Cell Dev Biol. 2021 May 13;9:671841. doi: 10.3389/fcell.2021.671841. eCollection 2021.
is a key gene during development, homeostasis and repair after injury. We previously reported a knock-in line (with the Cre-ERT2 cassette inserted in frame with the start codon of exon 1), called thereafter , to target FGF10 cells. While this line allowed fairly efficient and specific labeling of FGF10 cells during the embryonic stage, it failed to target these cells after birth, particularly in the postnatal lung, which has been the focus of our research. We report here the generation and validation of a new knock-in line (called thereafter ) with the insertion of the expression cassette in frame with the stop codon of exon 3. heterozygous mice exhibited comparable expression levels to wild type animals. However, a mismatch between and expression levels was observed in lungs. In addition, lung and limb agenesis were observed in homozygous embryos suggesting a loss of functional allele in mice. Bioinformatic analysis shows that the 3'UTR, where the Cre-ERT2 cassette is inserted, contains numerous putative transcription factor binding sites. By crossing this line with tdTomato reporter line, we demonstrated that tdTomato expression faithfully recapitulated expression during development. Importantly, mouse is capable of significantly targeting FGF10 cells in the adult lung. Therefore, despite the aforementioned limitations, this new line opens the way for future mechanistic experiments involving the postnatal lung.
是发育、体内稳态及损伤后修复过程中的关键基因。我们之前报道了一种敲入品系（Cre-ERT2盒插入到外显子1起始密码子的读框内），此后称为 ，用于靶向FGF10细胞。虽然该品系在胚胎期能相当高效且特异性地标记FGF10细胞，但出生后，尤其是在出生后的肺中（这一直是我们的研究重点），它无法靶向这些细胞。我们在此报告一种新的敲入品系（此后称为 ）的产生及验证，其表达盒插入到外显子3终止密码子的读框内。 杂合小鼠的 表达水平与野生型动物相当。然而，在 肺中观察到 与 表达水平不匹配。此外，在纯合胚胎中观察到肺和肢体发育不全，提示 小鼠中功能性等位基因缺失。生物信息学分析表明，Cre-ERT2盒插入的3'UTR包含众多假定的转录因子结合位点。通过将该品系与tdTomato报告基因品系杂交，我们证明tdTomato表达在发育过程中忠实地重现了 表达。重要的是， 小鼠能够在成年肺中显著靶向FGF10细胞。因此，尽管有上述局限性，这个新的 品系为未来涉及出生后肺的机制实验开辟了道路。
