Peyro Mohaddeseh, Dickson Andrew M, Mofrad Mohammad R K
Molecular Cell Biomechanics Laboratory, Departments of Bioengineering and Mechanical Engineering, University of California, Berkeley, CA 94720.
Molecular Biophysics and Integrative Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720.
Mol Biol Cell. 2021 Nov 1;32(21):ar31. doi: 10.1091/mbc.E21-04-0161. Epub 2021 Sep 2.
Nucleocytoplasmic traffic of nucleic acids and proteins across the nuclear envelop via the nuclear pore complexes (NPCs) is vital for eukaryotic cells. NPCs screen transported macromolecules based on their morphology and surface chemistry. This selective nature of the NPC-mediated traffic is essential for regulating the fundamental functions of the nucleus, such as gene regulation, protein synthesis, and mechanotransduction. Despite the fundamental role of the NPC in cell and nuclear biology, the detailed mechanisms underlying how the NPC works have remained largely unknown. The critical components of NPCs enabling their selective barrier function are the natively unfolded phenylalanine- and glycine-rich proteins called "FG-nucleoporins" (FG Nups). These intrinsically disordered proteins are tethered to the inner wall of the NPC, and together form a highly dynamic polymeric meshwork whose physicochemical conformation has been the subject of intense debate. We observed that specific sequence features (called largest positive like-charge regions, or lpLCRs), characterized by extended subsequences that only possess positively charged amino acids, significantly affect the conformation of FG Nups inside the NPC. Here we investigate how the presence of lpLCRs affects the interactions between FG Nups and their interactions with the cargo complex. We combine coarse-grained molecular dynamics simulations with time-resolved force distribution analysis to disordered proteins to explore the behavior of the system. Our results suggest that the number of charged residues in the lpLCR domain directly governs the average distance between Phe residues and the intensity of interaction between them. As a result, the number of charged residues within lpLCR determines the balance between the hydrophobic interaction and the electrostatic repulsion and governs how dense and disordered the hydrophobic network formed by FG Nups is. Moreover, changing the number of charged residues in an lpLCR domain can interfere with ultrafast and transient interactions between FG Nups and the cargo complex.
核酸和蛋白质通过核孔复合体（NPCs）在细胞核质之间的运输对于真核细胞至关重要。NPCs根据运输的大分子的形态和表面化学性质对其进行筛选。NPC介导的运输的这种选择性本质对于调节细胞核的基本功能，如基因调控、蛋白质合成和机械转导至关重要。尽管NPC在细胞和核生物学中具有重要作用，但其工作的详细机制在很大程度上仍然未知。使NPC具有选择性屏障功能的关键成分是天然未折叠的富含苯丙氨酸和甘氨酸的蛋白质，称为“FG核孔蛋白”（FG Nups）。这些内在无序的蛋白质附着在NPC的内壁上，共同形成一个高度动态的聚合物网络，其物理化学构象一直是激烈争论的主题。我们观察到特定的序列特征（称为最大正电荷相似区域，或lpLCRs），其特征是仅包含带正电荷氨基酸的延伸子序列，显著影响NPC内部FG Nups的构象。在这里，我们研究lpLCRs的存在如何影响FG Nups之间的相互作用以及它们与货物复合体的相互作用。我们将粗粒度分子动力学模拟与对无序蛋白质的时间分辨力分布分析相结合，以探索系统的行为。我们的结果表明，lpLCR结构域中带电残基的数量直接控制苯丙氨酸残基之间的平均距离及其相互作用的强度。因此，lpLCR内带电残基的数量决定了疏水相互作用和静电排斥之间的平衡，并控制由FG Nups形成的疏水网络的密度和无序程度。此外，改变lpLCR结构域中带电残基的数量会干扰FG Nups与货物复合体之间的超快和瞬时相互作用。
