The genetic dissection of seamless tube shape control in the Drosophila trachea

 描述（由申请人提供）：为了构建功能性血管网络，内皮管必须采用定型拓扑结构。在脊椎动物的血管中观察到多细胞和单细胞管拓扑结构。无缝管是单细胞的，没有连接。无缝管在脊椎动物血管网络内的多种情况下被发现。无缝管在这些情况下的功能作用知之甚少，因为无缝管形态发生的细胞机制知之甚少。然而，无缝管是保守的跨门，包括在呼吸（气管）系统的果蝇幼虫，其中强大的正向遗传方法可以用来研究无缝管形态发生。为了构建功能性血管网络，内皮管也必须采用定型的形状。内皮细胞分泌称为糖萼的管腔（顶端）细胞外基质（aECM），其由分泌的和膜结合的糖蛋白和蛋白聚糖组成。糖萼结构缺陷导致内皮管扩张缺陷。事实上，aECM在调节管生长中具有保守的作用，包括在果蝇气管管中。果蝇气管的无缝管分泌一种基于几丁质的aECM，称为角质层，其在无缝管形态发生中的作用尚不清楚。我们对个体分泌的基质因子组织成有序的aECM的分子途径了解不多。此外，aECM形成顶膜以调节管形态发生的分子途径是完全未知的。我将使用果蝇幼虫气管系统的无缝管作为一个模型来阐明新的遗传途径控制无缝管形状通过调节aECM的结构/功能。我已经确定了无缝管aECM在调节管形状和完整性方面的作用。缺乏几丁质酶的终末细胞表现出顶膜囊肿和不连续性。我还发现了一组新的突变体（水肿和哮喘），表型“几丁质生物合成”突变体。我假设这些突变体可能影响调节无缝管aECM的结构或功能的新途径。水肿和哮喘编码锌指转录因子（分别为CG 11966和zif），表明它们调节aECM组分的表达或靶向管腔。在具体目标1中，我将确定无缝管中aECM组织的遗传要求，包括测试已知位于Zif下游的保守极性因子在将aECM组分靶向管腔中的作用。在具体目标2中，我将确定无缝管aECM调节管形状的细胞和分子途径。我将测试的作用无缝管aECM在调节顶端肌动球蛋白组织使用遗传和创新的活细胞成像技术。

 DESCRIPTION (provided by applicant): To build a functional vascular network, endothelial tubes must adopt stereotyped topologies. Both multicellular and unicellular tube topologies are observed in vertebrate vasculatures. Seamless tubes are unicellular and unbound by junctions. Seamless tubes are found in multiple contexts within vertebrate vascular networks. The functional roles of seamless tubes in these contexts are poorly understood because the cellular mechanisms of seamless tube morphogenesis are poorly understood. Nevertheless, seamless tubes are conserved across phyla, including in the respiratory (tracheal) system of Drosophila melanogaster larvae, where powerful forward genetic approaches can be utilized to study seamless tube morphogenesis. To build a functional vascular network, endothelial tubes must also adopt a stereotyped shape. Endothelial cells secrete a luminal (apical) extracellular matrix (aECM) called the glycocalyx, comprised of secreted and membrane-bound glycoproteins and proteoglycans. Defects in glycocalyx structure lead to defects in endothelial tube expansion. Indeed, aECMs have a conserved role in regulating tube growth, including in Drosophila tracheal tubes. The seamless tubes of the Drosophila trachea secrete a chitin-based aECM called the cuticle, whose role in seamless tube morphogenesis not known. We have a poor understanding of molecular pathways through which individual secreted matrix factors are organized into an ordered aECM. Moreover, the molecular pathways through which aECMs shape the apical membrane to regulate tube morphogenesis are entirely unknown. I will use the seamless tubes of the Drosophila larval tracheal system as a model to elucidate novel genetic pathways controlling seamless tube shape by regulating aECM structure/function. I have identified a role for the seamless tube aECM in regulating tube shape and integrity. Chitin synthase-deficient terminal cells exhibit apical membrane cysts and discontinuities. I have also identified a set of novel mutants (ichor and asthmatic) that phenocopy "chitin biogenesis" mutants. I hypothesize that these mutants may affect novel pathway(s) regulating the structure or function of a seamless tube aECM. ichor and asthmatic encode zinc- finger transcription factors (CG11966 and zif, respectively), suggesting they regulate the expression or targeting of aECM components to the lumen. In Specific Aim 1, I will identify the genetic requirements for aECM organization in seamless tubes, including testing the role of a conserved polarity factor known to be downstream of Zif, in targeting aECM components to the lumen. In Specific Aim 2, I will identify cellular and molecular pathways through which the seamless tube aECM regulates tube shape. I will test a role for the seamless tube aECM in regulating apical actomyosin organization using both genetic and innovative live cell imaging techniques.