Real time dynamics of β-catenin expression during Hydra development, regeneration and Wnt signalling activation
Real time dynamics of β-catenin expression during Hydra development, regeneration and Wnt signalling activation
复制标题
DOI：
10.1387/ijdb.180092ct
复制
发表时间：
2018-01-01
影响因子：
0.7
通讯作者：
Tortiglione, Claudia
中科院分区：
文献类型：
--
作者：
Iachetta, Roberta;Ambrosone, Alfredo;Tortiglione, Claudia
了解推动形态发生和再生的动态细胞行为是生物学中的一个长期挑战。实时成像与基因编码的记者一起，可以提供必要的工具来解决这个问题，允许在体内监测在各种发育过程中感兴趣的基因的空间和时间表达动态。经典的Wnt/β-catenin信号控制着整个动物界在发育、再生和成年过程中过多的细胞活动。一些记者已经在动物模型中被制造出来，以揭示活跃的Wnt信号的位置。为了在体内监测Wnt/β-catenin信号通路的活性，我们构建了一个由Hydraβ-catenin启动子驱动的转Wnt/β-catenin信号通路的Hydra，利用光片荧光显微镜研究了Wnt/β-cat信号通路在萌发、再生和化学激活过程中的表达动态。对β-CAT-EGFP系的实时成像概括了先前报道的β-连环蛋白的内源表达模式，并揭示了Wnt/β-连环蛋白信号的新位点的动态出现，这些新的位点早期通过原位杂交的方法躲避了检测。通过将β-连环蛋白启动子的Wnt活性读出效率与先进的成像技术相结合，我们创建了一个新的模型系统来实时监测体内Hydraβ-CAT调控序列的活性，并为揭示许多其他生理环境中的β-连环蛋白调控开辟了道路。
Understanding the dynamic cellular behaviours driving morphogenesis and regeneration is a long-standing challenge in biology. Live imaging, together with genetically encoded reporters, may provide the necessary tool to address this issue, permitting the in vivo monitoring of the spatial and temporal expression dynamics of a gene of interest during a variety of developmental processes. Canonical Wnt/beta-catenin signalling controls a plethora of cellular activities during development, regeneration and adulthood throughout the animal kingdom. Several reporters have been produced in animal models to reveal sites of active Wnt signalling. In order to monitor in vivo Wnt/beta-catenin signalling activity in the freshwater polyp Hydra vulgaris, we generated a beta-cat-eGFP transgenic Hydra, in which eGFP is driven by the Hydra beta-catenin promoter.We characterized the expression dynamics during budding, regeneration and chemical activation of the Wnt/beta-cat signalling pathway using light sheet fluorescence microscopy. Live imaging of the beta-cat-eGFP lines recapitulated the previously reported endogenous expression pattern of beta-catenin and revealed the dynamic appearance of novel sites of Wnt/beta-catenin signalling, that earlier evaded detection by mean of in situ hybridization. By combining the Wnt activity read-out efficiency of the beta-catenin promoter with advanced imaging, we have created a novel model system to monitor in real time the activity of Hydra beta-cat regulatory sequences in vivo, and open the path to reveal beta-catenin modulation in many other physiological contexts.