Nuclear speckle liquid-liquid phase separation dynamics in senescence and aging

衰老和衰老过程中的核散斑液-液相分离动力学

• 批准号: 10604564
• 负责人: William Aaron Dion
• 金额: $ 4.77万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10604564
• 关键词: Affect; Age; Aging; Caenorhabditis elegans; Cell Aging; Cell Cycle; Cell model; Cells; ChIP-seq; Chromatin; Chronology; Core Protein; Couples; Data; Defect; Diffuse; Embryo; Endoplasmic Reticulum; Ensure; Environment; Evolution; Exhibits; Fibroblasts; Fluorescence Recovery After Photobleaching; Gene Expression; Genes; Genetic; Genetic Transcription; Health; Homeostasis; Hour; In Vitro; Inflammatory; Knowledge; Link; Liquid substance; Liver; Longevity; Mammalian Cell; Membrane; Methods; Modeling; Molecular; Morphology; Mus; N-terminal; Nuclear; Organelles; Organism; Orthologous Gene; Phase; Physical condensation; Physiological Processes; Probability; Process; Proteins; Proteome; Rejuvenation; Resistance; Role; SRSF2 gene; Scaffolding Protein; Signal Transduction; Son; System; Tamoxifen; Telomere Shortening; Testing; Time; Tissues; XBP1 gene; aged; cell injury; confocal imaging; druggable target; endoplasmic reticulum stress; fluidity; improved; in vivo; misfolded protein; model organism; mutant; novel; pharmacologic; protein aggregation; protein folding; proteostasis; response; senescence; transcriptome sequencing

Abstract Organismal health requires a consistent and balanced internal environment known as homeostasis. Different physiological processes maintain proper levels of biomolecules at a cellular level, and several of these mechanisms lose efficacy with age. Proteostasis, sustained levels of correctly folded proteins in the endoplasmic reticulum (ER), is maintained by the Unfolded Protein Response (UPR). Excessive misfolded proteins in the ER activate the three branches of the UPR, facilitating adaptive processes to restore a balanced proteome in the cell. Aging is associated with the loss of proteostasis and the accumulation of senescent cells – cells that no longer replicate and secrete pro-inflammatory signals – that exhibit a dysfunctional UPR. The molecular mechanisms underlying the altered UPR in senescent cells are unclear. We hypothesize that the liquid-liquid phase separation (LLPS) dynamics of a nuclear biomolecular condensate, the nuclear speckle (NS), link cellular senescence to the UPR. The 12-hour, XBP1s-dependent clock that functions independently of the 24-hour clock or the cell cycle establishes 12-hour ultradian rhythms of NS LLPS dynamics. These rhythms regulate NS morphology and fluidity through SON, the NS core protein. High SON levels create a diffuse, fluid NS and boost the expression of UPR-associated genes. In contrast, low SON levels result in a spherical, stagnant NS and a blunted expression of UPR genes. We have recently found that SON levels decrease, and that the NS becomes more spherical during cellular senescence. These data suggest that changes to NS LLPS dynamics are hallmarks of cellular senescence and aging. Here, we propose two aims to examine how NS LLPS dynamics change in vitro during cellular senescence and in vivo throughout chronological aging. In the first aim, we will use a mouse embryonic fibroblast line with a GFP-tagged NS that can be induced to enter senescence. This model will examine how established 12-hour rhythms of NS LLPS dynamics change during senescence and how restoring SON levels affects NS LLPS dynamics in senescent cells. We will also pharmacologically boost the diffuseness of the NS to determine if its fluidity can be increased during senescence. The second aim will use a Caenorhabditis elegans (C. elegans) model with a GFP-tagged NS. We will examine how NS LLPS dynamics change throughout aging and if genetic and pharmacological methods that make the NS more diffuse in mammalian cells can similarly affect NS LLPS dynamics in C. elegans and boost proteostasis in aged organisms. These aims will establish changes to NS LLPS dynamics as hallmarks of senescence and aging. Furthermore, we intend to show that NS LLPS dynamics is a druggable target and that therapies could return the NS morphology and fluidity to a pre-senescent state.

抽象的 有机体健康需要一个一致且平衡的内部环境，即稳态。不同的 生理过程在细胞水平上保持适当水平的生物分子，其中一些 机制随着年龄的增长而失去效率。蛋白质，在内质中持续的正确折叠蛋白水平 网状（ER）由未折叠的蛋白质反应（UPR）维持。急诊室中过多的不折叠蛋白 激活UPR的三个分支，支持自适应过程以恢复平衡的蛋白质组 细胞。衰老与蛋白抑制作用的丧失和感觉细胞的积累有关 - 无需 更长的复制和秘密的促炎信号 - 存在功能失调的UPR。分子 感觉细胞中UPR改变的机制尚不清楚。我们假设液体液体 核生物分子冷凝物，核斑点（NS），连接细胞的相分离（LLP）动力学 对UPR的感应。与24小时时钟无关的12小时XBP1依赖时钟 或细胞周期建立了NS LLPS动力学的12小时超级节奏。这些节奏调节ns NS核心蛋白儿子的形态和流动性。高儿子级别产生弥漫性，流体NS和增强 与UPR相关基因的表达。相反，低儿子水平导致球形，停滞的ns和a UPR基因的表达钝。我们最近发现儿子的水平下降，NS变为 细胞感应期间的球形更多。这些数据表明，NS LLPS动态的更改是 细胞感应和衰老的标志。在这里，我们提出了两个目的，以研究NS LLP的动态如何 整个年代老化过程中细胞感应和体内的体外变化。在第一个目标中，我们将 使用具有GFP标记的NS的小鼠胚胎成纤维细胞系，可引起进入感应。这 模型将检查NS LLPS动力学在感应过程中的12小时节奏以及如何如何变化 恢复儿子水平会影响感觉细胞中的NS LLPS动力学。我们还将在药品上提升 NS的扩散以确定在感应期间是否可以增加其流动性。第二个目标将使用 具有GFP标记的NS的秀丽隐杆线虫（秀丽隐杆线虫）模型。我们将研究NS LLP的动态 整个衰老的变化，以及是否使NS更加分散的遗传和药物方法 哺乳动物细胞类似地影响秀丽隐杆线虫中的NS LLPS动力学，并在老年生物体中增强蛋白质的抑制。 这些目标将建立对NS LLPS动力学的变化，作为感应和衰老的标志。此外， 我们打算表明NS LLPS动态是一个可吸毒的靶标，并且疗法可以返回NS 形态和流动性到感周期的状态。