Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
RNA. 2011 Jan;17(1):134-44. doi: 10.1261/rna.1210411. Epub 2010 Oct 29.
The biogenesis of a localization-competent mRNP begins in the nucleus. It is thought that the coordinated action of nuclear and cytoplasmic components of the localization machinery is required for the efficient export and subsequent subcellular localization of these mRNAs in the cytoplasm. Using quantitative poly(A)(+) and transcript-specific fluorescent in situ hybridization, we analyzed different nonessential nucleoporins and nuclear pore-associated proteins for their potential role in mRNA export and localization. We found that Nup60p, a nuclear pore protein located on the nucleoplasmic side of the nuclear pore complex, was required for the mRNA localization pathway. In a Δnup60 background, localized mRNAs were preferentially retained within the nucleus compared to nonlocalized transcripts. However, the export block was only partial and some transcripts could still reach the cytoplasm. Importantly, downstream processes were also affected. Localization of ASH1 and IST2 mRNAs to the bud was impaired in the Δnup60 background, suggesting that the assembly of a localization competent mRNP ("locasome") was inhibited when NUP60 was deleted. These results demonstrate transcript specificity of a nuclear mRNA retention defect and identify a specific nucleoporin as a functional component of the localization pathway in budding yeast.
定位能力的 mRNP 的生物发生始于细胞核。人们认为，定位机制的核和细胞质成分的协调作用对于这些 mRNA 在细胞质中的有效输出和随后的亚细胞定位是必需的。使用定量 poly(A)(+) 和转录特异性荧光原位杂交，我们分析了不同的非必需核孔蛋白和核孔相关蛋白在 mRNA 输出和定位中的潜在作用。我们发现，位于核孔复合物核质侧的核孔蛋白 Nup60p 对于 mRNA 定位途径是必需的。在 Δnup60 背景下，与非定位转录物相比，定位的 mRNAs 更倾向于保留在核内。然而，出口阻塞只是部分的，一些转录物仍然可以到达细胞质。重要的是，下游过程也受到影响。在 Δnup60 背景下，ASH1 和 IST2 mRNA 向芽的定位受损，表明当 NUP60 缺失时，组装具有定位能力的 mRNP（“locasome”）被抑制。这些结果证明了核 mRNA 保留缺陷的转录物特异性，并确定了一种特定的核孔蛋白作为芽殖酵母中定位途径的功能组成部分。
