Protein Phosphatase 1 Holoenzyme Formation

摘要 磷酸化是细胞中最普遍、最可逆的翻译后修饰之一。这些酶 负责控制细胞的磷酸化状态的是激酶，它催化细胞内 底物的γ-磷酸部分，以及催化逆水解反应的磷酸酶， 从磷酸化底物中去除磷酸盐部分。因此，磷酸酶动态逆转 激动酶的作用。因为磷酸化对所有生物过程都是至关重要的，从细胞生长到 分化到发育，激酶和磷酸酶相互作用的位置和持续时间 必须在细胞内的时间和空间上都进行精细的调节。因此，当这个紧身衣 调节被打乱，磷酸化信号的调节失调，其后果最常见的是 疾病。删除两个PP1调节剂-SDS22(PPP1R7)或抑制物-3(I3；PPP1R11或Ypi1)中的一个 在酵母中)-在酵母中是致命的(必需基因)，突出了它们的生物学意义。然而，由于他们的 自发现以来，SDS22和I3被分配了不同的生物学角色，包括在有丝分裂中的角色(SDS22)， E3连接酶功能(I3)、PP1生物发生等。因此，尽管很明显SDS22和I3是 必需的PP1调节剂，它们真正的生物学功能(S)，特别是它们的作用机制(S)仍然存在 未知。这阻碍了理解它们在PP1生物学中的作用的进展。在细胞中，这些蛋白质形成 异二聚体(SDS22：PP1和I3：PP1)和异三聚体(SDS22：I3：PP1；SIP)PP1复合体。这个 结构和功能(S)的个别二聚体复合体，如果和如何的结构和功能(S) 三聚体络合物不同于二聚体络合物以及每个络合物在PP1中的作用(S) 全酶的形成是该领域的主要问题。此外，更多的数据表明， Sip复合体需要AAA ATPase p37/p97。然而，SIP复合体解离的分子细节 也仍然难以捉摸。本研究项目使用了一种强大的综合方法， 将结构生物学与生化和细胞生物学实验相结合，以获得对 这些调节剂用来控制PP1活性和直接PP1全酶组装的分子机制。 由于pp1全酶在人类疾病中起着关键作用，这项拟议的工作将提供新的 靶向PP1全酶形成及其亚基选择性抑制PP1活性的策略 交流，这对于理解不同的购买力平价如何导致疾病至关重要。

ABSTRACT Phosphorylation is one of the most ubiquitous, reversible posttranslational modifications in cells. The enzymes responsible for controlling the phosphorylation state of the cell are kinases, which catalyze the transfer of the γ-phosphate moiety of ATP to substrates, and phosphatases, which catalyze the reverse hydrolysis reaction, the removal of the phosphate moiety from phosphorylated substrates. Thus, phosphatases dynamically reverse the effects of kinases. Because phosphorylation is critical for all biological processes from cell growth to differentiation to development, the location and duration of the reciprocal actions of kinases and phosphatases must be exquisitely regulated both temporally and spatially within the cell. Consequently, when this tight regulation is disrupted, dysregulation of phosphorylation signaling ensues and the consequence is most often disease. Deletion of either one of two PP1 regulators—SDS22 (PPP1R7) or Inhibitor-3 (I3; PPP1R11 or Ypi1 in yeast)—is lethal in yeast (essential genes), highlighting their biological significance. However, since their discovery, different biological roles have been assigned to SDS22 and I3, including roles in mitosis (SDS22), E3 ligase functionality (I3), PP1 biogenesis, among others. Thus, while it is clear that SDS22 and I3 are essential PP1 regulators, their true biological function(s) and especially their mechanism(s) of action are still unknown. This has hindered progress in understanding their roles in PP1 biology. In cells, these proteins form both heterodimeric (SDS22:PP1 and I3:PP1) and a heterotrimeric (SDS22:I3:PP1; SIP) PP1 complex. The structure and function(s) of the individual dimeric complexes, if and how the structure and function(s) of the trimeric complex differs from those of the dimeric complexes and the role(s) of each complex in PP1 holoenzyme formation are major questions in the field. Further, additional data suggest that dissociation of the SIP complex requires the AAA+ ATPase p37/p97. However, the molecular details of SIP complex dissociation have also remained elusive. The presented research project uses a powerful integrated approach that combines structural biology with biochemical and cell biology experiments to obtain novel insights into the molecular mechanisms used by these regulators to control PP1 activity and direct PP1 holoenzyme assembly. Because PP1 holoenzymes have critical roles in human diseases, the proposed work will provide novel strategies for selectively inhibiting PP1 activity by targeting the PP1 holoenzyme formation and subunit exchange, which is essential for understanding how distinct PPPs contribute to disease.