Department of Life Sciences, Ben-Gurion University, Beer-Sheva, Israel.
Mol Cell Biol. 2013 Nov;33(22):4371-80. doi: 10.1128/MCB.00713-13. Epub 2013 Sep 9.
The Drosophila melanogaster Spn-F, Ik2, and Javelin-like (Jvl) proteins interact to regulate oocyte mRNA localization and cytoskeleton organization. However, the mechanism by which these proteins interact remains unclear. Using antibodies to activated Ik2, we showed that this protein is found at the region of oocyte and follicle cell where microtubule minus ends are enriched. We demonstrate that germ line Ik2 activation is diminished both in jvl and in spn-F mutant ovaries. Structure-function analysis of Spn-F revealed that the C-terminal end is critical for protein function, since it alone was able to rescue spn-F sterility. On the other hand, germ line expression of Spn-F lacking its conserved C-terminal region (Spn-FΔC) phenocopied ik2, leading to production of ventralized eggshell and bicaudal embryos. In Spn-FΔC-expressing oocytes, Gurken protein is mislocalized and oskar mRNA and protein localization is disrupted. Expression of Ik2 rescued Spn-FΔC ovarian phenotypes. We found that whereas Spn-F physically interacts with Ik2 and Jvl, Spn-FΔC physically interacts with Ik2 but not with Jvl. Thus, expression of Spn-FΔC, which lacks the Jvl-interacting domain, probably interferes with interaction of Ik2 and Jvl. In summary, our results demonstrate that Spn-F mediates the interaction between Ik2 and Jvl to control Ik2 activity.
果蝇 Spn-F、Ik2 和 Javelin-like(Jvl)蛋白相互作用以调节卵母细胞 mRNA 的定位和细胞骨架的组织。然而,这些蛋白相互作用的机制尚不清楚。我们利用激活的 Ik2 抗体,证明该蛋白存在于卵母细胞和滤泡细胞中微管负端富集的区域。我们证实 jvl 和 spn-F 突变体的生殖系 Ik2 激活均减弱。Spn-F 的结构-功能分析表明,C 端对于蛋白功能至关重要,因为它本身就能挽救 spn-F 的不育性。另一方面,生殖系表达缺乏保守 C 端区域的 Spn-F(Spn-FΔC)表现出与 ik2 类似的表型,导致产生腹侧化卵壳和双尾胚胎。在 Spn-FΔC 表达的卵母细胞中,Gurken 蛋白发生定位错误,oskar mRNA 和蛋白的定位也受到干扰。Ik2 的表达挽救了 Spn-FΔC 的卵巢表型。我们发现 Spn-F 与 Ik2 和 Jvl 物理相互作用,而 Spn-FΔC 与 Ik2 物理相互作用,但不与 Jvl 相互作用。因此,表达缺乏与 Jvl 相互作用结构域的 Spn-FΔC,可能干扰了 Ik2 和 Jvl 的相互作用。总之,我们的结果表明,Spn-F 介导了 Ik2 和 Jvl 之间的相互作用,以控制 Ik2 的活性。