Webb Sarah E, Goulet Cécile, Chan Ching Man, Yuen Michael Y F, Miller Andrew L
Division of Life Science and State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science & Technology, Clear Water Bay, Kowloon, Hong Kong, People's Republic of China.
Division of Life Science, The Hong Kong University of Science & Technology, Clear Water Bay, Kowloon, Hong Kong, People's Republic of China.
Zygote. 2014 May;22(2):218-28. doi: 10.1017/S0967199413000051. Epub 2013 Mar 27.
The large and optically clear embryos of the zebrafish provide an excellent model system in which to study the dynamic assembly of the essential contractile band components, actin and myosin, via double fluorescent labelling in combination with confocal microscopy. We report the rapid appearance (i.e. within <2 min) of a restricted arc of F-actin patches along the prospective furrow plane in a central, apical region of the blastodisc cortex. These patches then fused with each other end-to-end forming multiple actin cables, which were subsequently bundled together forming an F-actin band. During this initial assembly phase, the F-actin-based structure did not elongate laterally, but was still restricted to an arc extending ~15° either side of the blastodisc apex. This initial assembly phase was then followed by an extension phase, where additional F-actin patches were added to each end of the original arc, thus extending it out to the edges of the blastodisc. The dynamics of phosphorylated myosin light chain 2 (MLC2) recruitment to this F-actin scaffold also reflect the two-phase nature of the contractile apparatus assembly. MLC2 was not associated with the initial F-actin arc, but MLC2 clusters were recruited and assembled into the extending ends of the band. We propose that the MLC2-free central region of the contractile apparatus acts to position and then extend the cleavage furrow in the correct plane, while the actomyosin ends alone generate the force required for furrow ingression. This biphasic assembly strategy may be required to successfully divide the early cells of large embryos.
斑马鱼的胚胎体积大且透明，是一个优秀的模型系统，可通过双荧光标记结合共聚焦显微镜来研究肌动蛋白和肌球蛋白等必需收缩带成分的动态组装。我们报告了在囊胚盘皮层中央顶端区域，沿着预期的沟平面快速出现（即<2分钟内）一条受限的F-肌动蛋白斑块弧。这些斑块随后端对端相互融合，形成多条肌动蛋白电缆，随后这些电缆聚集在一起形成一条F-肌动蛋白带。在这个初始组装阶段，基于F-肌动蛋白的结构没有横向伸长，而是仍然局限于在囊胚盘顶端两侧延伸约15°的弧。这个初始组装阶段之后是延伸阶段，在原始弧的两端添加额外的F-肌动蛋白斑块，从而将其延伸到囊胚盘的边缘。磷酸化肌球蛋白轻链2（MLC2）募集到这个F-肌动蛋白支架的动力学也反映了收缩装置组装的两阶段性质。MLC2与初始的F-肌动蛋白弧不相关，但MLC2簇被募集并组装到带的延伸端。我们提出，收缩装置中不含MLC2的中央区域起到定位作用，然后在正确的平面上延伸分裂沟，而仅肌动球蛋白末端产生沟侵入所需的力。这种双相组装策略可能是成功分裂大型胚胎早期细胞所必需的。
