The analysis of body axis formation in the common house spider Parasteatoda tepidariorum

背腹轴和前后轴的建立是两侧对称动物胚胎发育的基本过程。在蜘蛛胚胎中，一种叫做卵丘的细胞结构是胚胎轴的主要组织者。虽然其他节肢动物没有这样的结构，积云是非常相似的组织者，指定轴的两栖动物胚胎（Spemann组织者）。这两种组织者通过BMP信号传导发挥作用，并且能够在各自的胚胎中诱导次级体轴。积云的功能在普通的蜘蛛Parasteatoda tepidariorum中得到了最好的研究。已知的是，卵丘充当指定蜘蛛胚胎的背腹体轴所需的BMP信号的来源。积云产生于径向对称的胚盘中心，并向其外围迁移。通过这种迁移，积云打破了胚盘的径向对称性。然而，胚盘形成、卵丘特化和卵丘细胞中dpp活化的分子基础完全未知。在最初的研究计划中，我通过胚胎注射细胞骨架和RNA聚合酶抑制剂，并通过亲本RNA干扰Toll和EGF信号通路的候选基因以及许多其他候选转录因子，分析了胚盘的形成。虽然我可以证明胚盘的形成需要肌动蛋白（而不是微管蛋白）的细胞骨架和合子转录，候选基因的方法并没有导致胚盘或轴向图案表型。因此，更新建议的新方法是基于从新的转录组数据中选择的基因。一个试验性的RNAi筛选与10个新的候选基因已经导致两个显着的背腹图案表型和一个缺口基因样表型。这些结果表明，新的候选基因的选择是准确的，这是一个很好的方法来确定蜘蛛轴的决定最重要的基因。背腹表型特别有趣，因为其中一个基因是一种全新的卵丘迁移调节因子，另一个基因可能是BMP信号传导的上游（以前从未描述过）。鉴于这些令人兴奋的初步结果，从转录组数据中对剩余基因进行功能筛选可能会产生更多有趣和新颖的表型。

Setting up the dorsoventral and the anteroposterior body axes is a fundamental process during the embryonic development of all bilaterian animals. In spider embryos, a cellular structure called the cumulus is the main organizer of the embryonic axes. While other arthropods do not have such a structure, the cumulus is remarkably similar to the organizer that specifies the axes of amphibian embryos (Spemann organizer). Both organizers function via BMP signalling, and are able to induce a secondary body axis in the respective embryos. The function of the cumulus has been best studied in the common-house spider Parasteatoda tepidariorum. It is known that the cumulus acts as a source of BMP signalling required to specify the dorsoventral body axis of the spider embryo. The cumulus arises in the center of the radially-symmetric germ-disc and migrates to its periphery. Through this migration the cumulus breaks the radial symmetry of the germ-disc. However, the molecular basis of germ-disc formation, cumulus specification and dpp activation in the cells of the cumulus are completely unknown. In the initial research proposal, I analysed germ-disc formation via embryonic injection of cytoskeleton and RNA polymerase inhibitors, and via parental RNA interference with candidate genes of the Toll- and the EGF-signalling pathways as well as many other candidate transcription factors. While I could show that germ-disc formation requires the actin (but not the tubulin) cytoskeleton and zygotic transcription, the candidate gene approach did not result in germ-disc or axial patterning phenotypes. Therefore, the new approach of the renewal proposal is based on genes that have been selected from new transcriptome data. A pilot RNAi screen with ten new candidate genes already resulted in two striking dorsoventral patterning phenotypes and one gap-gene like phenotype. These results demonstrate that the selection of the new candidate genes is accurate, and that this is an excellent approach for identifying the genes most important for spider axis determination. The dorsoventral phenotypes are especially interesting, as one of the genes is a completely novel regulator of cumulus migration, and the other gene may be upstream of BMP signalling (something that has never been described before). Given these exciting preliminary results, a functional screen of the remaining genes from the transcriptome data will likely yield many more interesting and novel phenotypes.