Heinrich-Heine University Düsseldorf, Institute for Microbiology, Düsseldorf, Germany.
Mol Cell Proteomics. 2011 Dec;10(12):M111.011213. doi: 10.1074/mcp.M111.011213. Epub 2011 Aug 1.
Long-distance transport of mRNAs is crucial in determining spatio-temporal gene expression in eukaryotes. The RNA-binding protein Rrm4 constitutes a key component of microtubule-dependent mRNA transport in filaments of Ustilago maydis. Although a number of potential target mRNAs could be identified, cellular processes that depend on Rrm4-mediated transport remain largely unknown. Here, we used differential proteomics to show that ribosomal, mitochondrial, and cell wall-remodeling proteins, including the bacterial-type endochitinase Cts1, are differentially regulated in rrm4Δ filaments. In vivo UV crosslinking and immunoprecipitation and fluorescence in situ hybridization revealed that cts1 mRNA represents a direct target of Rrm4. Filaments of cts1Δ mutants aggregate in liquid culture suggesting an altered cell surface. In wild type cells Cts1 localizes predominantly at the growth cone, whereas it accumulates at both poles in rrm4Δ filaments. The endochitinase is secreted and associates most likely with the cell wall of filaments. Secretion is drastically impaired in filaments lacking Rrm4 or conventional kinesin Kin1 as well as in filaments with disrupted microtubules. Thus, Rrm4-mediated mRNA transport appears to be essential for efficient export of active Cts1, uncovering a novel molecular link between mRNA transport and the mechanism of secretion.
mRNA 的长距离运输对于真核生物中时空基因表达的决定至关重要。RNA 结合蛋白 Rrm4 构成了 Ustilago maydis 丝状体内微管依赖性 mRNA 运输的关键组成部分。尽管已经确定了许多潜在的靶 mRNA，但仍不清楚哪些细胞过程依赖于 Rrm4 介导的运输。在这里，我们使用差异蛋白质组学表明，核糖体、线粒体和细胞壁重塑蛋白，包括细菌型内切几丁质酶 Cts1，在 rrm4Δ 丝状体内的表达水平存在差异。体内 UV 交联和免疫沉淀以及荧光原位杂交显示，cts1 mRNA 是 Rrm4 的直接靶标。cts1Δ 突变体的丝状体细胞在液体培养中聚集，表明细胞表面发生了改变。在野生型细胞中，Cts1 主要定位于生长锥，而在 rrm4Δ 丝状体内则聚集在两个极上。内切几丁质酶被分泌出来，并可能与丝状体细胞壁结合。在缺乏 Rrm4 或传统的驱动蛋白 Kin1 的丝状体内以及在微管被破坏的丝状体内，分泌作用明显受损。因此，Rrm4 介导的 mRNA 运输对于活性 Cts1 的有效输出至关重要，揭示了 mRNA 运输与分泌机制之间的新的分子联系。
