Wong G K, Hoyle D H, Begg D A
Department of Anatomy and Cell Biology, Faculty of Medicine, University of Alberta, Edmonton, Canada.
Dev Biol. 1996 Nov 25;180(1):199-212. doi: 10.1006/dbio.1996.0295.
Spectrin has been implicated in a variety of different processes during late embryogenesis, after transcription of the zygotic genome has been activated. However, relatively little is known about the role of maternally derived spectrin during the early cleavage divisions that give rise to a multicellular embryo. To investigate the role of spectrin in early development, we have microinjected anti-spectrin antibodies into Patiria miniata starfish embryos to inhibit the activity of the maternal pool of spectrin. Microinjection of affinity-purified anti-spectrin antibody, or low to moderate doses of F(ab) fragments, into one blastomere of a two-cell-stage embryo caused a dose-dependent, progressive increase in the length of the cell cycle compared to the uninjected control blastomere. The progeny of injected blastomeres were unable to participate in the formation of a blastula epithelium, instead forming a loose aggregate of cells that eventually stopped dividing. When division stopped, the cells formed surface protrusions and became motile. At high doses of either whole antibody or F(ab) fragments, cells initiated, but failed to complete, cytokinesis. Blastomeres injected with high doses of F(ab) fragments also failed to reform nuclei and underwent variable periods of cell cycle arrest up to 12 hr. Injected embryos stained with BODIPY-phallacidin exhibited extensive disruption of the cortical actin cytoskeleton. These results support previous studies implicating spectrin in stabilizing the cell surface and maintaining the organization of the cortical cytoskeleton. They further suggest that spectrin is not required for the initiation or contraction of the cleavage furrow, but functions in the completion of cytokinesis. Most surprisingly, however, the results demonstrate that inhibition of spectrin function alters cell cycle timing, suggesting that disruption of the actin cytoskeleton inhibits progression through the cell cycle.
血影蛋白在合子基因组转录被激活后的胚胎后期发育过程中参与了多种不同的过程。然而，对于母源血影蛋白在产生多细胞胚胎的早期卵裂过程中的作用，人们了解得相对较少。为了研究血影蛋白在早期发育中的作用，我们将抗血影蛋白抗体显微注射到小红海盘车海星胚胎中，以抑制母源血影蛋白池的活性。将亲和纯化的抗血影蛋白抗体或低至中等剂量的F(ab)片段显微注射到二细胞期胚胎的一个卵裂球中，与未注射的对照卵裂球相比，导致细胞周期长度呈剂量依赖性逐渐增加。注射卵裂球的后代无法参与囊胚上皮的形成，而是形成松散的细胞聚集体，最终停止分裂。当分裂停止时，细胞形成表面突起并变得有运动能力。在高剂量的全抗体或F(ab)片段作用下，细胞启动但未能完成胞质分裂。注射高剂量F(ab)片段的卵裂球也未能重新形成细胞核，并经历长达12小时的不同时期的细胞周期停滞。用BODIPY-鬼笔环肽染色的注射胚胎显示皮质肌动蛋白细胞骨架受到广泛破坏。这些结果支持了先前的研究，即血影蛋白参与稳定细胞表面和维持皮质细胞骨架的组织。它们进一步表明，血影蛋白对于卵裂沟的启动或收缩不是必需的，而是在胞质分裂的完成中起作用。然而，最令人惊讶的是，结果表明血影蛋白功能的抑制改变了细胞周期时间，这表明肌动蛋白细胞骨架的破坏抑制了细胞周期的进程。
