Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK.
Trends Cell Biol. 2010 Jul;20(7):380-90. doi: 10.1016/j.tcb.2010.03.006. Epub 2010 May 3.
Localized mRNA provides spatial and temporal protein expression essential to cell development and physiology. To explore the mechanisms involved, considerable effort has been spent in establishing new and improved methods for visualizing mRNA. Here, we discuss how these techniques have extended our understanding of intracellular mRNA localization in a variety of organisms. In addition to increased ease and specificity of detection in fixed tissue, in situ hybridization methods now enable examination of mRNA distribution at the ultrastructural level with electron microscopy. Most significantly, methods for following the movement of mRNA in living cells are now in widespread use. These include the introduction of labeled transcripts by microinjection, hybridization based methods using labeled antisense probes and complementary transgenic methods for tagging endogenous mRNAs using bacteriophage components. These technical innovations are now being coupled with super-resolution light microscopy methods and promise to revolutionize our understanding of the dynamics and complexity of the molecular mechanism of mRNA localization.
局部化的 mRNA 提供了对细胞发育和生理至关重要的空间和时间蛋白质表达。为了探索所涉及的机制,人们花费了大量精力来建立新的和改进的方法来可视化 mRNA。在这里,我们讨论了这些技术如何扩展我们对各种生物体中细胞内 mRNA 定位的理解。除了在固定组织中提高检测的简便性和特异性外,原位杂交方法现在还可以通过电子显微镜检查超微结构水平的 mRNA 分布。最重要的是,现在广泛使用了跟踪活细胞中 mRNA 运动的方法。这些方法包括通过显微注射引入标记的转录本、使用标记的反义探针的杂交方法以及使用噬菌体成分标记内源性 mRNAs 的互补转基因方法。这些技术创新现在正在与超分辨率荧光显微镜方法相结合,并有望彻底改变我们对 mRNA 定位的分子机制的动力学和复杂性的理解。