Department of Biochemistry, University of Oxford, Oxford, United Kingdom.
PLoS One. 2013 Jul 23;8(7):e69978. doi: 10.1371/journal.pone.0069978. Print 2013.
In the Drosophila oocyte, gurken (grk) mRNA encodes a secreted TGF-α signal that specifies the future embryonic dorso-ventral axes by altering the fate of the surrounding epithelial follicle cells. We previously identified a number of RNA binding proteins that associate specifically with the 64 nucleotide grk localization signal, including the Drosophila orthologue of polypyrimidine tract-binding protein (PTB), Hephaestus (Heph). To test whether Heph is required for correct grk mRNA or protein function, we used immunoprecipitation to validate the association of Heph with grk mRNA and characterized the heph mutant phenotype. We found that Heph is a component of grk mRNP complexes but heph germline clones show that Heph is not required for grk mRNA localization. Instead, we identify a novel function for Heph in the germline and show that it is required for proper Grk protein localization. Furthermore, we show that Heph is required in the oocyte for the correct organization of the actin cytoskeleton and dorsal appendage morphogenesis. Our results highlight a requirement for an mRNA binding protein in the localization of Grk protein, which is independent of mRNA localization, and we propose that Heph is required in the germline for efficient Grk signalling to the somatic follicle cells during dorso-ventral patterning.
在果蝇卵母细胞中，gurken（grk）mRNA 编码一种分泌的 TGF-α 信号，通过改变周围上皮滤泡细胞的命运来指定未来的胚胎背腹轴。我们之前鉴定了许多与 grk 定位信号的 64 个核苷酸特异性结合的 RNA 结合蛋白，包括果蝇多嘧啶 tract-binding protein（PTB）的同源物 Hephaestus（Heph）。为了测试 Heph 是否需要正确的 grk mRNA 或蛋白功能，我们使用免疫沉淀来验证 Heph 与 grk mRNA 的结合，并对 heph 突变体表型进行了特征分析。我们发现 Heph 是 grk mRNP 复合物的一个组成部分，但 heph 生殖细胞克隆表明 Heph 对于 grk mRNA 的定位不是必需的。相反，我们在生殖细胞中发现了 Heph 的一个新功能，并表明它对于 Grk 蛋白的正确定位是必需的。此外，我们表明 Heph 在卵母细胞中对于肌动蛋白细胞骨架的正确组织和背附肢形态发生是必需的。我们的结果强调了在 grk 蛋白定位中需要一种 mRNA 结合蛋白，该蛋白与 mRNA 定位无关，并且我们提出 Heph 在生殖细胞中对于在背腹模式形成过程中向体细胞滤泡细胞进行有效的 Grk 信号转导是必需的。
