Schmitz Felizitas, Burtscher Ingo, Stauber Michael, Gossler Achim, Lickert Heiko
Institute of Diabetes and Regeneration Research, Helmholtz Zentrum München, Business Campus Garching, Parkring 11, Garching, D-85748, Germany.
Institute of Stem Cell Research, Helmholtz Zentrum München, Ingolstädter Landstraße 1, Neuherberg, D-85764, Germany.
Genesis. 2017 Nov;55(11). doi: 10.1002/dvg.23073. Epub 2017 Oct 13.
Cilia play a major role in the regulation of numerous signaling pathways and are essential for embryonic development. Mutations in genes affecting ciliary function can cause a variety of diseases in humans summarized as ciliopathies. To facilitate the detection and visualization of cilia in a temporal and spatial manner in mouse tissues, we generated a Cre-inducible cilium-specific reporter mouse line expressing an ARL13B-tRFP fusion protein driven by a CMV enhancer/chicken β actin promotor (pCAG) from the Hprt locus. We detected bright and specific ciliary signals by immunostainings of various mono- and multiciliated tissues and by time-lapse live-cell analysis of cultured embryos and organ explant cultures. Additionally, we monitored cilium assembly and disassembly in embryonic fibroblast cells using live-cell imaging. Thus, the ARL13B-tRFP reporter mouse strain is a valuable tool for the investigation of ciliary structure and function in a tissue-specific manner to understand processes, such as ciliary protein trafficking or cilium-dependent signaling in vitro and in vivo.
纤毛在众多信号通路的调节中起主要作用，对胚胎发育至关重要。影响纤毛功能的基因突变可导致人类多种疾病，统称为纤毛病。为了便于在小鼠组织中以时空方式检测和可视化纤毛，我们构建了一种Cre诱导的纤毛特异性报告基因小鼠品系，该品系从Hprt位点表达由CMV增强子/鸡β肌动蛋白启动子（pCAG）驱动的ARL13B-tRFP融合蛋白。我们通过对各种单纤毛和多纤毛组织进行免疫染色，以及对培养的胚胎和器官外植体培养物进行延时活细胞分析，检测到明亮且特异的纤毛信号。此外，我们使用活细胞成像监测胚胎成纤维细胞中的纤毛组装和拆卸。因此，ARL13B-tRFP报告基因小鼠品系是一种有价值的工具，可用于以组织特异性方式研究纤毛结构和功能，以了解体外和体内的纤毛蛋白运输或纤毛依赖性信号传导等过程。
