Characterisation of the co-adapter function of the E3 ligase Neuralized (Neur) during DSL-ligand induced Notch signalling

在所有后生动物中，缺口信号在许多发育过程中起着至关重要的作用。此外，通过参与成人干细胞的维持，它是包括人类在内的几个成人组织的动态平衡所必需的。完整的配体诱导的通路激活需要内吞作用。它是由膜相关的E3连接酶神经化(Neur)和Mindomb1(Mib1)启动的。它们与DSL配体的胞内区(ICD)结合，介导赖氨酸(K)侧链的泛素化(UBI)。泛素化的配体通过其泛素结合域被内吞适配液面(Lqf)/Epsin识别，从而连接到内吞核心机制。在果蝇中，这两个E3连接酶在翅膀发育过程中可以在功能上相互替代，表明它们可以执行类似的功能，尽管它们除了ubi所需的环(真正有趣的新基因)指状域(RF)外没有任何序列相似性。然而，Mib1不能取代Neur在神经发生过程中的功能，这表明Neur可以以不同于Mib1的方式启动Notch信号传递。我们最近的工作支持了这一观点，揭示了不能泛素化的配体可以被Nur激活，但不能被Mib1激活。因此，尽管Nur具有MIb1所不能提供的E3连接酶活性，但它还有另一种未知的功能。在这个应用中，我们将进一步描述Nur的功能，并遵循Nur是一种新型的内吞共适配子的假设，它与Lqf/Epsin形成复合体，以另一种方式将配体连接到核心内吞机制。

Notch signalling plays a crucial role in many developmental processes in all metazoans. Moreover, it is required for homeostasis of several adult tissues, including in humans, through its involvement in maintenance of adult stem cells. Endocytosis is required for the full ligand induced activation of the pathway. It is initiated by the membrane-associated E3 ligases Neuralized (Neur) and Mindbomb1 (Mib1). They bind to the intracellular domains (ICDs) of DSL ligands to mediate the ubiquitylation (ubi) on lysine (K) side chains. The ubiquitylated ligands are recognised by the endocytic adapter Liquid Facets (Lqf)/Epsin via its ubiquitin binding domains and thereby connected to the endocytic core machinery. In Drosophila, the two E3 ligases can functionally substitute for one another during wing development, indicating that they can perform similar functions, even though they share no sequence similarity apart from a RING (Really Interesting New Gene) Finger domain (RF), which is required for ubi. However, Mib1 cannot replace the function of Neur during neurogenesis, suggesting that Neur can initiate Notch signalling in a manner different from Mib1. Our recent work supports this notion by revealing that ligands that cannot be ubiquitylated can be activated by Neur, but not Mib1. Hence, Neur has another so far unknown function despite its E3 ligase activity, which cannot be provided by Mib1. In this application, we will further characterise the function of Neur and follow the hypothesis that Neur is a novel type of endocytic co-adapter that forms a complex with Lqf/Epsin to connect the ligands in an additional way to the core endocytic machinery.