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

基本信息

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
项目类别：
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

项目摘要

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患病器官和组织的恢复。人类、蝾螈具有快速再生器官的非凡能力，包括四肢、脊髓、我们的目标是发现这些动物如何在几周内重建功能性的成年组织。从发育到退化——我们器官的健康和功能取决于产生然而，我们目前对再生的理解主要基于研究。 mRNA 而非直接评估最终修复所需的蛋白质。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，我们还缺乏以相同规模和分辨率研究 mRNA 翻译成蛋白质的工具。墨西哥蝾螈与其他蝾螈一样，以其终生“青春永驻”而闻名。通过结合尖端方法，截肢后重新生长整个肢体的能力尚不完全清楚。在翻译研究中，我们能够证明，与哺乳动物不同，蝾螈的严重损伤令人惊讶的是，当细胞增殖很少时，蛋白质合成就会快速激活。不寻常的分子反应是再生脊椎动物特有的特征，并且依赖于此外，我们发现哺乳动物雷帕霉素靶点 (mTOR) 通路的比例少于 20%。 axolotl mRNA 在任何给定时间都会被翻译，其余部分在翻译之外以“自由”状态存在我们将检验蝾螈中的“自由”转录本可能在空间上组织成的假设。无膜区室由功能相关且翻译共同调节的 mRNA 组成对细胞生存和细胞命运规范至关重要的转录本在这些区室和细胞之间穿梭我们进一步确定了核糖体对促进伤口愈合和再生的控制。再生时高度依赖于蝾螈超越应激激活信号的能力相反，我们将测试结构/序列的假设。 Axolotl mTOR 成分的具体差异可以揭示上游调节的功能差异物种之间的蛋白质合成以及将“应激反应”信号重新利用到生物体的巨大能力这些发现表明哺乳动物的愈合不良可能是由于“生长和再生”信号造成的。由于损伤部位存在明显的细胞信号反应，而不是由于再生能力的固有缺乏最后，我们发现蝾螈的肢体截肢会触发一些基因的选择性翻译。核糖体蛋白质而不是其他蛋白质，与蛋白质合成的“爆发”一致，因此我们将测试大胆的。假设蝾螈可能组装特殊核糖体的不同子集以促进选择性表达该提案旨在提供对伤口愈合和再生至关重要的转录本。关于为什么有些物种可以再生而另一些物种不能再生的机械理解。