Local Synthesis in the Mauthner Axon

Mauthner 轴突的局部合成

• 批准号: 9010251
• 负责人: Edward Koenig
• 金额: $ 22.19万
• 依托单位: SUNY at Buffalo
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-09-15 至 1994-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9010251&HistoricalAwards=false
• 关键词: Local; Synthesis; Mauthner; Axon

Axoplasm, located in the axons of nerve cells, is composed of cytoskeletal and cytosolic proteins which are supplied to the axon by a poorly understood process of slow transport. Since all proteins exhibit a characteristic of half-life, it is likely that during slow transport in the axon, proteins undergo significant biological decay through degradation. In order to maintain the steady state of protein composition and mass of the axon due to significant losses through biological decay, local mechanisms for the replenishment of proteins are necessary. This research project will examine the localization and distribution of selected cytoskeletal protein manufacturing systems and ascertain what proteins are synthesized locally in the axon. Although the evidence available indicates that myelinated axons have a limited capacity for local synthesis and that local requirements for newly synthesized proteins in a short half-life period are probably modest, the amount of synthesis occurring over the whole axon is probably significant. Experiments will be conducted at the cellular level on the Mauthner neuron, the M-axon, an identifiable cell in the CNS of the goldfish with a large myelinated axon extending the length of the spinal cord. A major advantage in using the M-axon preparation is that the myelin sheath is separated from glial cell bodies, making it possible to separate the axon from the myelin sheath uncontaminated for the analysis of the cell body and the axon. Specialized methods, designed and developed in Dr. Koenig's laboratory, for studying ultrapure microscopic samples will be used. The investigator will utilize in situ hybridization techniques to localize ribosomal RNA in the M-axon and to assess its density and distribution. Messenger RNAs will be localized in the M-axon for actin and/or tubulin, important cytoskeletal proteins that are supplied to the axon from the cell body by slow transport. Analysis of axonal polypeptides synthesized in the M-axon, in an in vitro preparation, will be performed in order to characterize which proteins are synthesized locally in the short half-life period and to establish that labeled proteins are due to de novo synthesis. Proteins synthesized in the M-cell axon in vivo will be compared with those synthesized in vitro in order to identify proteins synthesized under physiological conditions, and proteins with a longer half-life. The results of this study will address an important unresolved issue of how the mature axon compensates for biological decay of its exogenously supplied proteins.

轴浆位于神经细胞的轴突中，由 细胞骨架蛋白和胞质蛋白，它们被提供给 轴突通过一个鲜为人知的缓慢运输过程。 由于所有 蛋白质表现出半衰期的特征，很可能， 在轴突的缓慢运输过程中，蛋白质经历了显著的 生物通过降解而腐烂。 为了保持 稳定状态的蛋白质组成和质量的轴突，由于 通过生物腐烂造成的重大损失， 蛋白质的补充是必要的。 本研究 项目将研究本地化和分布 选择细胞骨架蛋白质制造系统，并确定 哪些蛋白质是在轴突中局部合成的 虽然 现有的证据表明，有髓鞘的轴突具有有限的 地方综合能力和地方对 在短半衰期内新合成的蛋白质， 可能是适度的，在整个过程中发生的合成量 axon轴突is probably大概is significant重要. 实验将在 在Mauthner神经元、M-轴突、 在金鱼的中枢神经系统中， 延伸脊髓长度的有髓轴突。 一个主要 使用M-轴突制剂的优点是髓磷脂 鞘与神经胶质细胞体分离，使得有可能 将轴突与未被污染的髓鞘分离， 细胞体和轴突的分析。 专门的方法， 在Koenig博士的实验室设计和开发， 将使用超纯显微样品。 研究者 将利用原位杂交技术 核糖体RNA在M-轴突，并评估其密度， 分布 信使RNA将定位在M-轴突中， 肌动蛋白和/或微管蛋白，重要的细胞骨架蛋白， 通过缓慢运输从细胞体供应到轴突。 在一个人的M-轴突中合成的轴突多肽的分析， 将进行体外制备，以表征 哪些蛋白质在短半衰期内局部合成 期间，并建立标记的蛋白质是由于从头 合成. 在体内M细胞轴突中合成的蛋白质将 与体外合成的那些进行比较，以鉴定 在生理条件下合成的蛋白质， 半衰期更长。 这项研究的结果将解决 一个重要的未解决的问题， 其外源蛋白质的生物衰变。