Exploring function and dynamics in the human VCP/p97-cofactor network

P97(又称VCP或CDC48)是一种泛素导向的AAA+型ATPase，具有从应激反应途径、蛋白质降解到细胞内信号转导等多种不同的细胞活动。它们对基因组稳定性和蛋白质动态平衡至关重要，与退行性疾病和衰老有关。为了实现这些功能，p97被认为与其他辅因子蛋白结合，并形成途径特异性的p97-辅因子复合体。然而，在大多数情况下，这些复合体只是定义得很模糊，而-相反-大量预测的辅因子没有被分配到功能复合体。在之前的资助期间，我们成功地使用应激诱导的辅因子定位筛选结合功能验证在一个p97介导的过程中定义了一个新的辅因子复合体，即内溶酶体损伤反应。我们的目标是现在进一步开发和系统地在一系列细胞应激反应途径中应用该方法，以在功能上分配p97辅因子并定义不同的途径特异性p97复合体。该方法将得到基于细胞敲除和RNAi方法的功能研究的补充，使用一系列已建立的读数，以及辅因子复合体组装的生化和微观分析。这将有助于创建p97辅因子系统的全面图景，允许比较p97介导的细胞应激反应，并研究它们在扰动下的关系。预期的结果将填补对已出现的多样化和深远的应激反应系统的理解的主要差距，并有望揭示可能的治疗干预策略，用于p97相关和相关的退行性疾病。

P97 (also called VCP or Cdc48) is a ubiquitin-directed AAA+-type ATPase with a large number of diverse cellular activities that range from stress response pathways and protein degradation to intracellular signalling. They are critical for genomic stability and protein homeostasis with relevance for degenerative disease and aging. To fulfil these functions, p97 is believed to associate with alternative sets of cofactor proteins and form pathway-specific p97-cofactor complexes. However, in most cases, these complexes are only poorly defined, while - conversely - a large number of predicted cofactors have not been assigned to functional complexes.In the previous funding period, we have successfully used stress-induced cofactor localisation screening combined with functional validation to define a novel cofactor complex in one p97-mediated process, the endo-lysosomal damage response. Our goal is to now further develop and apply the approach systematically in a range of cellular stress response pathways to functionally assign p97 cofactors and define distinct pathway-specific p97 complexes. The approach will be complemented with functional studies based on cellular knock-out and RNAi approaches using a range of established readouts, as well as with biochemical and microscopic analyses of cofactor complex assembly. This will help create a comprehensive picture of the p97-cofactor system, allow comparison of p97-mediated cellular stress responses and to study their relationship upon perturbations.The expected results will close a major gap in the understanding of what has emerged as a diverse and far-reaching stress response system, and will hopefully also reveal possible strategies for therapeutic intervention in p97-associated and related degenerative disorders.