JNK regulation of desmosomes in development.

描述（由申请人提供）：在发育过程中，表皮保护胚胎免受化学、生物和机械应力的影响。这个器官在提供张力方面也是必不可少的，这是形成器官所必需的，同时也允许形态发生所必需的细胞形状变化。表皮的完整性是由细胞间连接复合物（包括桥粒）介导的。然而，对于桥粒在胚胎发育过程中是如何调控的，我们所知相对较少。我的实验室发现了c-Jun NH(2)-末端激酶（JNK）信号在调节非洲爪蟾蝌蚪表皮的桥粒蛋白中的潜在新作用。JNK信号已被证明在胚胎发生过程中调节多种基因的转录或直接磷酸化细胞骨架蛋白。但是，这个MAP激酶家族成员从未被证明调节桥粒体功能。我们发现，JNK信号传导减少导致的表型与人类的情况一致，这种情况导致桥粒功能缺陷，如表皮脆性和冒泡，以及心脏和颅面缺陷。此外，在JNK靶点的蛋白质组学筛选中，我们发现了两个蛋白；与哺乳动物桥粒相关的外周蛋白和外周蛋白。本研究旨在结合多种方法研究表皮中JNK信号功能的时空缺失。接下来将描述在整个胚胎和细胞水平上的影响。我们将使用反义技术、光激活和移植试验来进行表皮特异性JNK1功能丧失。桥粒将通过电子和共聚焦显微镜进行分析。最后，我们的目标是确定JNK是否通过转录或使用定量PCR和共免疫沉淀的直接磷酸化间接调节桥粒蛋白，epiplakin和periplakin。我相信这是一项创新的资助，它使用全胚胎方法，连接发育信号和桥粒体功能。

DESCRIPTION (provided by applicant): During development the epidermis protects the embryo from chemical, biological and mechanical stresses. This organ is also essential in providing tension, necessary for shaping organs while also allowing for cell shape changes necessary for morphogenesis. The integrity of the epidermis is mediated by intercellular junctional complexes, including desmosomes. However, relatively little is known about how desmosomes are regulated during embryonic development. My lab has uncovered a potentially new role for c-Jun NH(2)- terminal kinase (JNK) signaling in regulating desmosomal proteins in the epidermis of Xenopus laevis tadpoles. JNK signaling has been shown to regulate transcription of a variety of genes or to directly phosphorylate cytoskeletal proteins during embryogenesis. But, this MAP kinase family member has never been shown to regulate desmosomal function. We have found that decreased JNK signaling results phenotypes consistent with human conditions resulting from defective desmosomal function such as epidermal fragility and bubbling as well as heart and craniofacial defects. Further, in a proteomics screen for JNK targets we uncovered two plakins; epiplakin and periplakin that are associated with desmosomes in mammals. This proposal aims to use a combination of methods to perform temporal and spatial loss of function of JNK signaling in the epidermis. This will be followed by characterizing the effects in the whole embryo and at the cellular level. We will perform epidermal specific loss of JNK1 function using antisense technology, photoactivation and transplant assays. Desmosomes will be analyzed by electron and confocal microscopy. Finally we aim to determine whether JNK regulates the desmosomal proteins, epiplakin and periplakin, indirectly via transcription or by direct phosphorylation using quantitative PCR and co-immunoprecipitations. I believe this to be an innovative grant that uses whole embryo approaches and bridges developmental signaling and desmosomal function.