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: 10674724
• 负责人: Maria Barna
• 金额: $ 34.36万
• 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/10674724
• 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; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; Health; Heart; Human; Injury; Knock-in; 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; cell fate specification; genome-wide; healing; human tissue; limb amputation; limb regeneration; mRNA Translation; mouse model; novel; organ regeneration; 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疾病器官和组织的修复。不像 在人类中，火蜥蜴具有迅速再生器官的非凡能力，包括四肢、脊髓、 心和脑。我们的目标是在几周内发现这些动物如何重建具有功能的成人组织。 从发育到退化-我们器官的健康和功能取决于 适当的组织特异性蛋白质。然而，我们目前对再生的理解很大程度上是基于研究 而不是直接评估最终修复所需的蛋白质。这部分归因于 技术限制-微阵列和RNA-Seq技术彻底改变了我们对转录的理解- 但直到最近，我们还缺乏在相同规模和分辨率下研究信使核糖核酸转化为蛋白质的工具。 墨西哥的紫杉醇以其终生的“青春”而闻名。Aaxolotls与其他火蜥蜴分享令人惊讶的 并不完全了解截肢后再生整个肢体的能力。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，轴突的严重损伤 令人惊讶的是，在细胞几乎没有增殖的时候，蛋白质合成却能迅速激活。这 不寻常的分子反应是再生脊椎动物特有的特征，依赖于 哺乳动物雷帕霉素(MTOR)途径的靶点。此外，我们发现，显著不到20%的所有人 Axolot1的mRNAs在任何给定的时间被翻译，其余的在翻译之外以自由的状态存在 机械设备。我们将检验这样一种假设，即Axolot1中的“自由”转录可能在空间上组织成 由功能相关和翻译共调控的mRNAs和 对细胞存活和细胞命运决定至关重要的转录本在这些隔间和 核糖体促进伤口愈合和再生。我们进一步确认了蛋白质合成的控制 在再生时间高度依赖于Axolotl超过应力激活信号的能力 而是促进mTOR通路的激活。我们将测试结构/序列的假设 Axolotl mTOR组分的具体差异可以揭示上游调控的功能差异 不同物种之间蛋白质合成的差异，以及将“应激反应”信号重新定位为 “生长和再生”的信号。这些发现表明，哺乳动物的愈合不良可能是 由于损伤部位有明显的细胞信号反应，而不是由于天生缺乏再生能力 潜力。最后，我们发现，截肢在Axolotl触发选择性翻译的一些 核糖体蛋白质，而不是其他蛋白质，与蛋白质合成的“爆发”相一致。因此，我们将测试大胆的 关于Axolotls可能组装不同亚群的特化核糖体以促进选择性表达的假设 对伤口愈合和再生至关重要的文字记录。总而言之，这项提案试图提供一部小说 机械地理解为什么一些物种可以再生，而另一些物种不能。