Rapid remodeling of the translatome underlying wound healing and regeneration

伤口愈合和再生中翻译组的快速重塑

• 批准号: 10445695
• 负责人: Maria Barna
• 金额: $ 34.2万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10445695
• 关键词: Adult; Ambystoma; Ambystoma mexicanum; Amputation; Animals; Binding Sites; Brain; Cell Fate Control; Cell Proliferation; Cell Survival; Cells; Complex; Cytoplasmic Granules; Development; Digit structure; Disease; Evolution; Exhibits; FRAP1 gene; Foundations; Genetic Transcription; Genetic Translation; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; Health; Heart; Human; Injury; Knock-in; Light; Limb structure; Lysosomes; Mammals; Membrane; Messenger RNA; Methods; Molecular; Mus; Natural regeneration; Organ; Organism; Pathway interactions; Phosphotransferases; Polyribosomes; Production; Protein Biosynthesis; Proteins; RNA, Messenger, Stored; Regulation; Resolution; Rest; Ribosomal Proteins; Ribosomes; Role; Salamander; Science; Signal Transduction; Site; Spinal Cord; Stress; Stress Response Signaling; Structure; Technology; Testing; Time; Tissues; Transcript; Translating; Translational Activation; Translational Regulation; Translational Research; Translations; Vertebrates; base; cell fate specification; genome-wide; healing; human tissue; limb amputation; limb regeneration; mouse model; novel; polysome profiling; protein activation; recruit; regeneration potential; regenerative; repaired; response; restoration; ribosome profiling; selective expression; severe injury; tissue regeneration; tool; transcriptome; transcriptome sequencing; translational impact; translatome; wound healing

The biggest biomedical challenge of this century is the restoration of diseased organs and tissues. Unlike humans, salamanders have the extraordinary ability to rapidly regenerate organs, including limbs, spinal cords, hearts and brains. Our goal is to discover how these animals rebuild functional adult tissues in a matter of weeks. From development through degeneration – the health and function of our organs depends on production of appropriate tissue-specific proteins. Yet, our current understanding of regeneration is largely based on studies of mRNA and not on direct assessment of proteins that are ultimately required for repair. This is in part due to technical limitations – microarray and RNA-Seq technologies revolutionized our understanding of transcription- but until recently we lacked the tools to study translation of mRNA into protein at the same scale and resolution. The Mexican axolotl is famous for its lifelong “youthfulness”. Axolotls share with other salamanders the surprising and incompletely understood ability to regrow entire limbs after amputation. By combining cutting-edge methods in translation research, we were able to demonstrate that, unlike in mammals, severe injury in the axolotl surprisingly results in rapid activation of protein synthesis at a time when there is little cellular proliferation. This unusual molecular response is a feature specific to regenerative vertebrates and relies on activation of the mammalian target of rapamycin (mTOR) pathway. Moreover, we find that remarkably fewer than 20% of all axolotl mRNAs are translated at any given time, the remainder exist in a ‘free’ state outside the translation machinery. We will test the hypothesis that the ‘free’ transcripts in the axolotl may be spatially organized into membrane-less compartments comprised of functionally-related and translationally co-regulated mRNAs and that transcripts critical for cell survival and cell fate specification shuttle between these compartments and the ribosome to facilitate wound healing and regeneration. We have further identified that control of protein synthesis at the time of regeneration is highly dependent on the ability of the Axolotl to surpass a stress activating signal and instead promote activation of the mTOR pathway. We will test the hypothesis that the structural/sequence specific differences in Axolotl mTOR components can shed light on functional differences in upstream regulation of protein synthesis between species and the remarkably ability to repurpose a ‘stress-response’ signal to a ‘growth and regeneration’ signal. These findings suggest the possibility that poor healing in mammals may be due to a distinct cellular signaling response at the site of injury rather than to an inherent lack of regenerative potential. Lastly, we have found that amputation of the limb in the axolotl triggers selective translation of some ribosomal proteins but not others, coincident with the “burst” in protein synthesis. We will therefore test the bold hypothesis that axolotls may assemble distinct subsets of specialized ribosomes to facilitate selective expression of transcripts critical for wound healing and regeneration. Together, this proposal seeks to provide a novel mechanistic understanding as to why some species can regenerate while others cannot.

本世纪最大的生物医学挑战是恢复患病的器官和组织。与众不同 人类，萨拉曼人具有非凡的能力，可以快速再生器官，包括四肢，脊髓， 心和大脑。我们的目标是发现这些动物在几周内如何重建功能性成人组织。 从发展到退化 - 我们的器官的健康和功能取决于生产 适当的组织特异性蛋白。但是，我们目前对再生的理解主要基于研究 mRNA而不是直接评估最终修复所需的蛋白质。这部分是由于 技术局限性 - 微阵列和RNA-Seq技术彻底改变了我们对转录的理解 - 但是直到最近，我们还缺乏研究mRNA翻译成蛋白质的工具。 墨西哥Axolotl以其终生的“年轻人”而闻名。 Axolotls与其他Salamanders共享惊喜 截肢后不完全理解的是改革整个四肢。通过结合尖端方法 在翻译研究中，我们能够证明，与哺乳动物不同，Axolotl严重损伤 令人惊讶的是，在几乎没有细胞增殖的时候，蛋白质合成的快速激活。这 异常的分子反应是再生脊椎动物的特征，并且依赖于激活 雷帕霉素（MTOR）途径的哺乳动物靶标。此外，我们发现全部少于20％ Axolotl mRNA在任何给定时间都会翻译，其余的都存在于翻译以外的“自由”状态 机械。我们将检验以下假设：axolotl中的“自由”笔录可以在空间上组织 与功能相关和翻译共同调节的mRNA的无膜隔室和 这些转录物对于这些隔室与细胞脂肪规格班车至关重要 核糖体可促进伤口愈合和再生。我们进一步确定了蛋白质合成的控制 在再生时 而是促进MTOR途径的激活。我们将测试结构/序列的假设 Axolotl MTOR组件的具体差异可以阐明上游调节的功能差异 物种之间的蛋白质合成与将“应力反应”信号重现为A的蛋白质合成 “生长和再生”信号。这些发现表明，哺乳动物的愈合不良的可能性可能是 由于损伤部位有明显的细胞信号反应，而不是继承缺乏再生 潜在的。最后，我们发现Axolotl触发器中肢体的截肢选择性翻译 核糖体蛋白质，而不是其他蛋白质，与蛋白质合成中的“爆发”一致。因此，我们将测试大胆 假设Axolotls可能会组装不同的专用核糖体的不同子集以促进选择性表达 转录物对于伤口愈合和再生至关重要。这项提议共同寻求提供小说 关于为什么有些物种可以再生而而另一些物种不能进行的机械理解。