In vivo Bedeutung des MAPK-Signalweg-Inhibitors SPRED - Charakterisierung der Funktion durch "knockout"-Modelle

本项目的目的是阐明SPRED蛋白在体外和完整生物体中的多功能和复杂功能。根据我们之前的工作，我们打算：阐明与强迫症样行为相关的SPRED2 KO小鼠应激激素分泌升高的机制。我们打算研究KO小鼠杏仁核突触的行为、电生理特性和解剖特征。通过生成SPRED1和-2.3的诱导型或组织特异性敲除模型，规避SPRED1蛋白被其他家族成员明显的功能代偿和双敲除的胚胎致死性。为了验证SPRED2与微管蛋白的蛋白/蛋白相互作用，这是通过下拉试验和随后的质谱分析确定的。在分子水平上，我们想研究SPRED2是否可能调节微管蛋白聚合，或者SPRED2是否可能通过功能参与调节微管上的囊泡运输。这种相互作用可能是生物学基本过程的一部分，因此可能与我们对真核细胞细胞内运输的理解高度相关。动物模型提供了在整个生物体的生理背景下重新评估分子观察的可能性。

Aim of this project is to elucidate the versatile and complex functions of SPRED proteins in vitro but also in an intact organism. Based on our previous work, we intend:1. To clarify the mechanism responsible for the elevated stress hormone production in SPRED2 KO mice, which is associated with OCD-like behaviour. We intend to investigate the behaviour, electrophysiological properties of synapses in the amygdala, and anatomical characteristics of KO mice.2. To circumvent the obvious functional compensation of SPRED proteins by other family members and the embryonic lethality of double knockouts through generation of inducible or tissue-specific kncokou models for SPRED1 and -2.3. To verify the protein/protein interaction of SPRED2 with Tubulins, which was identified by pull-down assays and subsequent mass spectrometrical analyses. On the molecular level, we would like to investigate if SPRED2 might regulate Tubulin polymerisation or if SPRED2 might by functionally involved in the regulation of vesicle transport along microtubules. This interaction may be part of a fundamental process in biology and may, therefore, be of high relevance for our understanding of intracellular transport in eukaryotic cells.The animal models offer the possibilty to reassess the molecular observations in a physiological context in entire organism.