Gard D L, Cha B J, Roeder A D
Department of Biology, University of Utah, Salt Lake City 84112, USA.
Zygote. 1995 Feb;3(1):17-26. doi: 10.1017/s0967199400002331.
We used confocal immunofluorescence microscopy to examine spindle migration, morphology and orientation during the maturation of Xenopus oocytes, in the presence or absence of cytochalasin B (CB), an inhibitor of actin assembly. Treatment with CB during maturation (10-50 micrograms/ml beginning 0-3 h prior to addition of progesterone) disrupted the normal organisation of the novel MTOC and transient microtubule array (MTOC-TMA complex) that serves as the immediate precursor of the first meiotic spindle, suggesting that F-actin plays an important role in the assembly or maintenance of this complex. However, CB treatment did not block translocation of the MTOC-TMA complex to the oocyte cortex, suggesting that MTOC-TMA translocation is not dependent on an actin-based mechanism. Bipolar spindles were observed in CB-treated oocytes fixed during both M1 and M2. However, rotation of the M1 and M2 spindles into an orientation orthogonal to the oocyte surface was inhibited by CB. Rhodamine-phalloidin revealed a concentration of F-actin at the site of M1 spindle attachment, further suggesting that cortical actin is required for anchoring and rotation of the meiotic spindles. Finally, the incidence of M1 monasters was significantly increased in CB-treated oocytes, suggesting that interactions between the nascent M1 spindle and cortex are dependent on F-actin.
我们使用共聚焦免疫荧光显微镜来检测非洲爪蟾卵母细胞成熟过程中纺锤体的迁移、形态和方向，实验条件为存在或不存在细胞松弛素B（CB），一种肌动蛋白组装抑制剂。在成熟过程中用CB处理（在添加孕酮前0 - 3小时开始，浓度为10 - 50微克/毫升）会破坏新型微管组织中心（MTOC）和瞬时微管阵列（MTOC - TMA复合体）的正常组织，该复合体是第一次减数分裂纺锤体的直接前体，这表明F - 肌动蛋白在该复合体的组装或维持中起重要作用。然而，CB处理并未阻止MTOC - TMA复合体向卵母细胞皮层的转运，这表明MTOC - TMA转运不依赖于基于肌动蛋白的机制。在M1和M2期固定的经CB处理的卵母细胞中观察到了双极纺锤体。然而，CB抑制了M1和M2纺锤体旋转到与卵母细胞表面正交的方向。罗丹明 - 鬼笔环肽显示在M1纺锤体附着部位有F - 肌动蛋白聚集，进一步表明皮层肌动蛋白是减数分裂纺锤体锚定和旋转所必需的。最后，在经CB处理的卵母细胞中，M1单星体的发生率显著增加，这表明新生M1纺锤体与皮层之间的相互作用依赖于F - 肌动蛋白。
