Deciphering molecular functions of the “multitalented” scaffold protein USH1G/SANS in primary cilia and the nucleus

SANS（scaffold protein containing ankyrin repeats and SAM domain）由USH 1G基因编码，是Usher综合征（Usher syndrome，USH）相关蛋白质相互作用组的核心支架分子。虽然USH 1G/SANS的缺陷主要导致患者的眼睛和内耳功能障碍，但它几乎无处不在。先前的研究已经证明，支架蛋白参与不同的细胞模块和过程，如机械敏感信号复合物的组装，初级纤毛发生，细胞内运输，内吞作用和前体mRNA剪接。在这里，我们将集中在分子间的相互作用和相关的SANS在初级纤毛和细胞核的动态过程。在我们的项目中，我们的目标是通过综合方法，结合生物化学，分子细胞生物学和生物物理学的不同方法以及分子建模，阐明USH 1G/SANS在初级纤毛和细胞核中的分子功能。我们将破译与配体蛋白的相互作用和它们的结合动力学和性质SANS，监测MLO的分子动力学和它们之间的活细胞中，和LLPS性质的MLO的两个细胞器，即初级纤毛基地和核斑点或Cajal体在核中。我们的具体目标如下：（1）揭示多价蛋白与SANS结合的物理化学基础。(2)阐明SANS在细胞核和初级纤毛分子动力学中的作用。(3)核和初级纤毛之间SANS依赖的分子互连的评价。(4)解析SANS在LLPS形成聚合物缩合物中的作用。通过我们的实验方法和计算机模拟，我们的目标是全面了解SANS在蛋白质-蛋白质相互作用和LLPS中作用的分子机制。我们不仅期待新的见解共同的机制和动力学和LLPS的相互作用，而且还进一步了解的剪接体和纤毛发生和/或deciliation的调节。此外，前瞻性结果将阐明核和睫状体MLO之间的联系。此外，我们希望查明LLP形成和“MLOar间”交换的变化在多大程度上与疾病相关，从而成为人类Usher综合征病理生理学的基础。

SANS (scaffold protein containing ankyrin repeats and SAM domain) is encoded by USH1G gene and serves as a central scaffold molecule of the protein interactome related to the human Usher syndrome (USH), the most common form of heritetery combined deaf-blindness. Although defects in USH1G/SANS predominately lead to dysfunctions in the eye and the inner ear in patients, it is almost ubiquitously expressed. Previous studies have demonstrated that the scaffold protein participates in diverse cell modules and processes, such as the assembly of mechanosensitive signaling complexes, primary ciliogenesis, intracellular transport, endocytosis, and pre-mRNA splicing. Here, we will focus on molecular interactions and the dynamics of processes related to SANS in primary cilia and the nucleus. In our project, we aim to unravel the molecular function of USH1G/SANS in primary cilia and the nucleus by integrative approaches, combining different methods from biochemistry, molecular cell biology and biophysics, as well as molecular modeling. We will decipher the interactions with ligand proteins and their binding kinetics and properties to SANS, monitor the molecular dynamics of MLOs and between them in the living cell, and the LLPS properties in the MLOs of both cell organelles, namely the primary ciliary base and the nuclear speckles or Cajal bodies in the nucleus. We have the following specific objectives: (1) Unraveling the physico-chemical basis for multivalent protein binding to SANS. (2) Elucidation the function of SANS for the molecular dynamics in the nucleus and primary cilia. (3) Evaluation of SANS-dependent molecular interconnections between nucleus and primary cilia. (4) Deciphering the role of SANS in the formation of polymer condensates by LLPS. With our experimental approaches and the in-silico modeling we aim to deliver a comprehensive view of the molecular mechanisms underlying the role of SANS in protein-protein interaction and LLPS. We expect not only novel insights into common mechanisms and the dynamics and interplay of LLPS, but also further understanding of the regulation of the spliceosome and ciliogenesis and /or deciliation. In addition, prospective results will elucidate the link between nuclear and ciliary MLOs. Moreover, we expect to pinpoint to what extent alterations in the formation of LLPS and in the “interMLOar” exchange are disease-relevant, underlying the pathophysiology of human Usher syndrome.