Motility screen identifies Drosophila IGF-II mRNA-binding protein--zipcode-binding protein acting in oogenesis and synaptogenesis.
Motility screen identifies Drosophila IGF-II mRNA-binding protein--zipcode-binding protein acting in oogenesis and synaptogenesis.
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DOI：
10.1371/journal.pgen.0040036
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发表时间：
2008-02
期刊：
影响因子：
4.5
通讯作者：
Hays TS
中科院分区：
文献类型：
--
作者：
Boylan KL;Mische S;Li M;Marqués G;Morin X;Chia W;Hays TS
特定mrna的定位可以建立局部蛋白质梯度，从而产生和控制细胞不对称的发展。虽然所有的证据都强调了细胞骨架在rna转运和定位中的重要性，但我们对这些事件是如何调节的了解有限。通过对GFP蛋白诱捕系中运动蛋白的视觉筛选，我们鉴定出果蝇IGF-II mrna结合蛋白（Imp），它是爪蟾Vg1 RNA结合蛋白和鸡邮政编码结合蛋白的同源物。在果蝇中，Imp是一个大型rna敏感复合体的一部分，该复合体在两种极化细胞类型中富集，即发育中的卵母细胞和神经元。使用延时共聚焦显微镜，我们确定动力蛋白和动力蛋白都有助于GFP-Imp颗粒的运输，并且卵室中的运输调节似乎与神经元中的运输调节不同。在果蝇中，功能缺失的Imp突变是合子致死性的，突变体在成年后死亡较晚。Imp在果蝇的卵发生过程中具有非必要的功能，但在胚胎发生和后期发育过程中具有必要的功能。Imp突变的种系克隆不会阻断母体mRNA定位或卵母细胞发育，但特定Imp异构体的过表达会破坏背侧/腹侧极性。我们在这里报道了功能缺失的Imp突变，以及Imp过表达，可以改变突触末端的生长。我们的数据显示Imp被转运到神经肌肉连接处，在那里它可能调节mRNA靶点的翻译。在卵母细胞中，Imp功能不是必需的，我们暗示了一个特定的Imp结构域在建立背腹极性。信使RNA的定位是蛋白质活动产生局部不对称的主要机制，并在多种生物功能中得到利用。mRNA定位和由此产生的蛋白质梯度对于胚胎轴的建立、细胞和神经元的极化运动以及突触信号的调节至关重要。目前，我们对mrna的组装、运输和定位所需的许多交易因素的了解还很初级。在这项研究中，我们利用体内运动试验来筛选活果蝇卵腔中主动运输的RNP复合物的成分。其中鉴定的果蝇IGF-II mRNA结合蛋白（Imp）是鸡邮政编码结合蛋白或人IGF-II mRNA结合蛋白的同源物。人IGF-II mRNA结合蛋白与乳腺肿瘤中癌细胞的转移行为有关，但其机制尚不清楚。我们证明，果蝇Imp RNP复合物在卵发生过程中，以及在神经元中，通过微管马达、动力蛋白和运动蛋白积极运输。我们发现在卵母细胞和神经元中转运的调节是不同的，并首次报道了果蝇Imp影响神经肌肉突触末端的生长。
The localization of specific mRNAs can establish local protein gradients that generate and control the development of cellular asymmetries. While all evidence underscores the importance of the cytoskeleton in the transport and localization of RNAs, we have limited knowledge of how these events are regulated. Using a visual screen for motile proteins in a collection of GFP protein trap lines, we identified the Drosophila IGF-II mRNA-binding protein (Imp), an ortholog of Xenopus Vg1 RNA binding protein and chicken zipcode-binding protein. In Drosophila, Imp is part of a large, RNase-sensitive complex that is enriched in two polarized cell types, the developing oocyte and the neuron. Using time-lapse confocal microscopy, we establish that both dynein and kinesin contribute to the transport of GFP-Imp particles, and that regulation of transport in egg chambers appears to differ from that in neurons. In Drosophila, loss-of-function Imp mutations are zygotic lethal, and mutants die late as pharate adults. Imp has a function in Drosophila oogenesis that is not essential, as well as functions that are essential during embryogenesis and later development. Germline clones of Imp mutations do not block maternal mRNA localization or oocyte development, but overexpression of a specific Imp isoform disrupts dorsal/ventral polarity. We report here that loss-of-function Imp mutations, as well as Imp overexpression, can alter synaptic terminal growth. Our data show that Imp is transported to the neuromuscular junction, where it may modulate the translation of mRNA targets. In oocytes, where Imp function is not essential, we implicate a specific Imp domain in the establishment of dorsoventral polarity. The localization of messenger RNA is a major mechanism to generate local asymmetries in protein activities and is utilized in a diverse array of biological functions. mRNA localization and the resultant protein gradients are critical for the establishment of embryonic axes, the polarized motility of cells and neurons, and the modulation of synaptic signaling. Presently, our knowledge of the many transacting factors required for the assembly, transport, and localization of mRNAs is rudimentary. In this study, we capitalize on an in vivo motility assay to screen for components of actively transported RNP complexes in live Drosophila egg chambers. One of the components identified, Drosophila IGF-II mRNA binding protein or Imp, is the homolog of chicken zipcode binding protein or human IGF-II mRNA binding protein. The human IGF-II mRNA binding protein is linked to the metastatic behavior of carcinoma cells in mammary tumors, but the mechanism is unclear. We demonstrate that the Drosophila Imp RNP complex, is actively transported in oogenesis, as well as in neurons by the microtubule motors, dynein and kinesin. We show that the regulation of transport is distinct in oocytes and neurons and report for the first time, that Drosophila Imp impacts growth of the neuromuscular synaptic terminal.
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