Schatten G, Simerly C, Asai D J, Szöke E, Cooke P, Schatten H
Integrated Microscopy Resource for Biomedical Research, University of Wisconsin, Madison 53706.
Dev Biol. 1988 Nov;130(1):74-86. doi: 10.1016/0012-1606(88)90415-0.
alpha-Tubulin in the microtubules of mouse oocytes and embryos is acetylated in a specific spatial and temporal sequence. In the unfertilized oocyte, a monoclonal antibody to the acetylated form of alpha-tubulin is bound predominantly at the poles of the arrested metaphase meiotic spindle. The labeling intensity of the spindle microtubules is weaker as observed by immunofluorescence using oocytes double-labeled for total tubulin and acetylated alpha-tubulin, and as measured by immuno high-voltage electron microscopy (immunoHVEM) with colloidal gold; cytasters are not acetylated. At meiotic anaphase, the spindle becomes labeled, and by telophase and during second polar body formation only the meiotic midbody is acetylated. The sperm axoneme retains its acetylation after incorporation though the interphase microtubules are not detected. First mitosis follows a pattern similar to that observed at the second meiosis and during interphase only the mitotic midbodies are acetylated. After treatment with cold, colcemid, or griseofulvin, the remaining stable microtubules are acetylated, but immunoHVEM observations suggest that these fibers might not have been acetylated prior to microtubule disruption. Taxol stabilization does not alter acetylation patterns. Acetylated microtubules are not necessarily old microtubules since acetylated fibers are observed at 30 sec after cold recovery. These results show the presence of acetylated microtubules during meiosis and mitosis and demonstrate a cell-cycle-specific pattern of acetylation, with acetylated microtubules found at the centrosomes at metaphase, an increase in spindle labeling at anaphase, and the selective deacetylation of all but midbody microtubules at telophase.
小鼠卵母细胞和胚胎微管中的α-微管蛋白以特定的空间和时间顺序被乙酰化。在未受精的卵母细胞中,针对α-微管蛋白乙酰化形式的单克隆抗体主要结合在减数分裂中期停滞纺锤体的两极。通过对总微管蛋白和乙酰化α-微管蛋白进行双重标记的卵母细胞进行免疫荧光观察,以及用胶体金免疫高压电子显微镜(immunoHVEM)测量,纺锤体微管的标记强度较弱;星体未被乙酰化。在减数分裂后期,纺锤体开始被标记,到末期以及第二极体形成期间,只有减数分裂中间体被乙酰化。精子轴丝在并入后仍保留其乙酰化状态,尽管未检测到间期微管。第一次有丝分裂遵循与第二次减数分裂相似的模式,在间期只有有丝分裂中间体被乙酰化。用冷、秋水仙酰胺或灰黄霉素处理后,剩余的稳定微管被乙酰化,但immunoHVEM观察表明,这些纤维在微管破坏之前可能未被乙酰化。紫杉醇稳定化不会改变乙酰化模式。乙酰化微管不一定是旧的微管,因为在冷恢复后30秒就观察到了乙酰化纤维。这些结果表明在减数分裂和有丝分裂过程中存在乙酰化微管,并证明了一种细胞周期特异性的乙酰化模式,在中期着丝粒处发现乙酰化微管,后期纺锤体标记增加,并在末期除中间体微管外所有微管的选择性去乙酰化。