Characterization of an RNA granule from developing brain
Characterization of an RNA granule from developing brain
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DOI：
10.1074/mcp.m500255-mcp200
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发表时间：
2006-04-01
影响因子：
7
通讯作者：
Sossin, WS
中科院分区：
文献类型：
--
作者：
Elvira, G;Wasiak, S;Sossin, WS
在大脑中，信使核糖核酸（mRNAs）从细胞体运输到突起，使得在远离细胞核的部位进行局部蛋白质翻译成为可能。通过亚细胞分级分离，我们从大鼠胚胎第18天的大脑中分离出一个组分，该组分富含类似无定形核糖体集合的结构。这个组分中编码β - 肌动蛋白的信使核糖核酸含量丰富，β - 肌动蛋白的信使核糖核酸在发育中的神经元的树突和轴突中运输。通过串联质谱法确定的该组分中丰富的蛋白质成分包括核糖体蛋白、RNA结合蛋白、微管相关蛋白（包括动力蛋白 dynein）以及一些仅被描述为潜在开放阅读框的蛋白质。RNA结合蛋白、被运输的信使核糖核酸、核糖体机制以及运输动力蛋白的结合将这些结构定义为RNA颗粒。在培养的海马神经元中表达所鉴定蛋白质的一个子集证实，蛋白质组学中鉴定的蛋白质存在于与核糖体和信使核糖核酸相关的神经突中。此外，许多表达的蛋白质共定位在一起。延时视频显微镜观察表明，含有其中一种蛋白质（DEAD盒3解旋酶）的复合物在海马神经元树突中迁移的速度与报道的RNA颗粒的速度相同。尽管颗粒的速度不受活动或神经营养因子脑源性神经营养因子（BDNF）的影响，但脑源性神经营养因子（而非活动）增加了移动颗粒的比例。这些研究确定了发育中的大脑中表达的RNA颗粒的分离方法和组成成分。
In brain, mRNAs are transported from the cell body to the processes, allowing for local protein translation at sites distant from the nucleus. Using subcellular fractionation, we isolated a fraction from rat embryonic day 18 brains enriched for structures that resemble amorphous collections of ribosomes. This fraction was enriched for the mRNA encoding beta-actin, an mRNA that is transported in dendrites and axons of developing neurons. Abundant protein components of this fraction, determined by tandem mass spectrometry, include ribosomal proteins, RNA-binding proteins, microtubule-associated proteins (including the motor protein dynein), and several proteins described only as potential open reading frames. The conjunction of RNA-binding proteins, transported mRNA, ribosomal machinery, and transporting motor proteins defines these structures as RNA granules. Expression of a subset of the identified proteins in cultured hippocampal neurons confirmed that proteins identified in the proteomics were present in neurites associated with ribosomes and mRNAs. Moreover many of the expressed proteins co-localized together. Time lapse video microscopy indicated that complexes containing one of these proteins, the DEAD box 3 helicase, migrated in dendrites of hippocampal neurons at the same speed as that reported for RNA granules. Although the speed of the granules was unchanged by activity or the neurotrophin brain-derived neurotrophic factor, brain-derived neurotrophic factor, but not activity, increased the proportion of moving granules. These studies define the isolation and composition of RNA granules expressed in developing brain.