Wulfkuhle J D, Petersen N S, Otto J J
Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47907-1392, USA.
Cell Motil Cytoskeleton. 1998;40(2):119-32. doi: 10.1002/(SICI)1097-0169(1998)40:2<119::AID-CM2>3.0.CO;2-A.
Drosophila neurosensory bristle development provides an excellent model system to study the role of the actin-based cytoskeleton in polarized cell growth. We used confocal fluorescence microscopy of isolated thoracic tissue to characterize changes in F-actin that occurred during macrochaete development in wild type flies and mutants that have aberrant bristle morphology. At the earliest stages in wild type bristle development, cortical patches of F-actin were present, but no bundles were observed. Actin bundles began to form at 31% of pupal development and became more prominent as development progressed. The F-actin patches gradually disappeared and were no longer present by 38% of pupal development. The distribution of F-actin in singed3 mutant macrochaetae was indistinguishable from wild type bristles until 35% of development when the actin bundles began to splay and appear ribbon-like. In forked36a bristles, the mutant phenotype was evident at earlier stages of development than the singed3 mutant. Wild type tissue stained with antibodies against the forked protein demonstrated that the forked protein colocalized with F-actin structures found in early and late stage developing macrochaetae. Antibodies against the singed protein showed it appeared to localize with F-actin structures only at later stages in development. These data suggested that the forked gene product was required for the initiation of fiber bundle formation and the singed gene product was required for the maintenance of fiber bundle morphology during bristle development. Similar analyses of singed3/forked36a double mutants provided additional genetic evidence that the forked gene product was required before the singed gene product. Further, the analyses suggested that at least one additional crosslinking protein was present in these bundles.
果蝇神经感觉刚毛的发育为研究基于肌动蛋白的细胞骨架在极化细胞生长中的作用提供了一个极佳的模型系统。我们使用分离的胸部组织的共聚焦荧光显微镜来表征野生型果蝇和具有异常刚毛形态的突变体在大刚毛发育过程中发生的F-肌动蛋白变化。在野生型刚毛发育的最早阶段，存在F-肌动蛋白的皮质斑块，但未观察到束状结构。肌动蛋白束在蛹发育的31% 时开始形成，并随着发育的进行变得更加明显。F-肌动蛋白斑块逐渐消失，到蛹发育的38% 时不再存在。在单毛3突变体大刚毛中，F-肌动蛋白的分布与野生型刚毛没有区别，直到发育的35% 时，肌动蛋白束开始散开并呈现带状。在叉状36a刚毛中，突变表型在发育的早期阶段比单毛3突变体更明显。用抗叉状蛋白的抗体对野生型组织进行染色表明，叉状蛋白与在发育中的大刚毛早期和晚期发现的F-肌动蛋白结构共定位。抗单毛蛋白的抗体显示，它似乎仅在发育的后期阶段与F-肌动蛋白结构定位。这些数据表明，叉状基因产物是纤维束形成起始所必需的，而单毛基因产物是刚毛发育过程中纤维束形态维持所必需的。对单毛3/叉状36a双突变体的类似分析提供了额外的遗传证据，表明叉状基因产物在单毛基因产物之前是必需的。此外，分析表明这些束中至少存在一种额外的交联蛋白。
