STRUCTURAL ANALYSIS OF THE NFATC CYTOPLASMIC TO NUCLEAR TRANSLOCATION MECHANISM

NFATC 细胞质核易位机制的结构分析

• 批准号: 8362176
• 负责人: Alys A Peisley
• 金额: $ 0.03万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2012-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8362176
• 关键词: Adopted; Binding; Biological Process; Calcineurin; Cell Nucleus; Complex; Data; Disease; Family member; Funding; Genetic Transcription; Grant; Importins; Mediating; N-terminal; National Center for Research Resources; Nuclear; Nuclear Export; Nuclear Import; Nuclear Translocation; Principal Investigator; Process; Protein Dephosphorylation; Protein Export Pathway; Proteins; Radiation; Reporting; Research; Research Infrastructure; Resources; Roentgen Rays; Serine; Signal Transduction; Source; Structure; United States National Institutes of Health; calcineurin phosphatase; cost; nuclear factors of activated T-cells; receptor; structural biology; transcription factor

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Many biological processes are exquisitely sensitive to the level of nuclear occupancy of NFATc proteins. The process of rapid nuclear import and export is regulated by an uncharacterized allosteric switch in the N-terminus of the protein, which regulates the alternate interaction between NFATc proteins and the nuclear import and export machinery. NFAT signaling is initiated by sustained low amplitude Ca2+ signals that activate the heterodimeric phosphatase calcineurin. Calcineurin binds directly to NFATc proteins through a conserved motif and dephosphorylates serines within the SP repeats and serine rich motifs (SRR and SP-repeats) in the N-terminus of NFATc family members. This dephosphorylation unmasks the nuclear localization sequence, allowing interaction with the importin complex and rapid cytoplasmic-to-nuclear translocation. In the nucleus NFAT proteins combine with other transcription factors (NFATn) to form specific transcription complexes. Dyrk1a and GSK3 then act sequentially to mediate export of the proteins from the nucleus. Rephosphorylation in the nucleus induces another conformational change, which exposes the nuclear export sequence (NES) and allows interaction with the nuclear export receptor Crm1. The mechanism of this allosteric switch in the N-terminus has not been defined structurally. The N-terminal region is predicted to be unfolded, while NMR spectra and SAXS data reveal a partially disordered protein. A number of intrinsically disordered regions in eukaryotic transcription factors have been reported to adopt structure upon binding to their partners. We are investigating NFAT in complex with a variety of binding partners in both the phosphorylated (cytoplasmic) and dephosphorylated (nuclear) forms to characterize the conformational change that occurs in the N-terminus of the protein by Small Angle X-ray Scattering.

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 许多生物过程对NFATc蛋白的核占有水平非常敏感。快速核输入和输出的过程是由蛋白质的N-末端中的未表征的变构开关调节的，其调节NFATc蛋白与核输入和输出机制之间的交替相互作用。NFAT信号传导由激活异二聚体磷酸酶钙调磷酸酶的持续低振幅Ca 2+信号启动。钙调磷酸酶通过保守基序直接结合NFATc蛋白，并使NFATc家族成员的N末端SP重复序列和富含丝氨酸的基序（SRR和SP重复序列）内的丝氨酸脱磷酸化。这种去磷酸化暴露了核定位序列，允许与importin复合物相互作用和快速的胞质到核易位。在细胞核中，NFAT蛋白质联合收割机与其他转录因子（NFATn）结合形成特异性转录复合物。然后Dyrk 1a和GSK 3依次作用以介导蛋白质从细胞核的输出。细胞核中的再磷酸化诱导另一种构象变化，其暴露核输出序列（内斯）并允许与核输出受体Crm 1相互作用。在N-末端的这种变构开关的机制尚未在结构上定义。N-末端区域被预测为未折叠的，而NMR光谱和SAXS数据揭示了部分无序的蛋白质。据报道，真核生物转录因子中的许多固有无序区域在与它们的配偶体结合时采用结构。我们正在研究NFAT与磷酸化（细胞质）和去磷酸化（细胞核）形式的各种结合伴侣的复合物，以通过小角X射线散射表征蛋白质N末端发生的构象变化。