Wolf K W, Joshi H C
Institut für Anthropologie, Johannes Gutenberg-Universität Mainz, Germany.
Mol Reprod Dev. 1996 Dec;45(4):547-59. doi: 10.1002/(SICI)1098-2795(199612)45:4<547::AID-MRD17>3.0.CO;2-T.
Meiotic spindles in males of higher Lepidotera are unusual in that the bulk of the spindle microtubules (MTs) ends about halfway between the equatorial plate and the centrosomes in metaphase. It appears worthwhile to determine how the MTs are nucleated, while their pole proximal ends are distant from the centrosomes. To this end, spermatocytes of Phragmatobia fuliginosa (Arctiidae), collected in the field, were double-labeled with antibodies to beta- and gamma-tubulin. The former antibody reveals the entire microtubular cytoskeleton, and the latter is directed against a newly-discovered tublin isoform that is prevalent in microtubule-organizing centers (MTOCs). The immunocytochemical work was supplemented by a fine structural analysis of MTOCs and spindles. Gamma-tubulin was clearly detected at the spindle poles, and prominent microtubular asters originated from these sites. Additionally, MT arrays at both sides of the equatorial plate in metaphase spermatocytes contained gamma-tubulin. The staining persisted in late anaphase, when kinetochore MTs are depolymerized. This indicates that at least nonkinetochore MTs contain gamma-tubulin. The analysis of ultrathin sections through spindles revealed large amounts of pericentriolar material at the spindles poles, in prometaphase through anaphase. The spindle MTs appeared as regular, straight elements in longitudinal sections. We assume that gamma-tubulin is located at the pole proximal ends of the MTs and/or is associated with the spindle MTs throughout their lengths. In order to distinguish between these possibilities, testes of Ephestia kuehniella (Pyralidae), a laboratory species, were cold-treated prior to double-labeling with antibodies to beta- and gamma-tubulin. The treatment was expected to depolymerize MTs. Astral MTs, which were nucleated end-on by gamma-tubulin-containing material, indeed depolymerized. In contrast, the gamma-tubulin-containing spindle MTs persisted. It is, therefore, conceivable that gamma-tubulin is associated with MTs throughout their lengths in male meiosis of Lepidoptera species. It is plausible that this association stabilizes the MTs against cold-induced disassembly.
高等鳞翅目雄性减数分裂纺锤体不同寻常之处在于，在中期时，纺锤体微管（MTs）的大部分末端位于赤道板和中心体之间大约一半的位置。确定微管如何成核似乎很有必要，因为它们靠近极的末端距离中心体较远。为此，对在野外采集的亮灰蝶（灯蛾科）精母细胞用抗β - 微管蛋白和γ - 微管蛋白的抗体进行双重标记。前一种抗体可显示整个微管细胞骨架，而后一种抗体针对的是一种新发现的在微管组织中心（MTOC）中普遍存在的微管蛋白异构体。免疫细胞化学工作辅以对微管组织中心和纺锤体的精细结构分析。在纺锤体极明显检测到γ - 微管蛋白，并且有明显的微管星体从这些位点发出。此外，中期精母细胞赤道板两侧的微管阵列含有γ - 微管蛋白。当动粒微管解聚时，这种染色在后期仍然存在。这表明至少非动粒微管含有γ - 微管蛋白。对纺锤体超薄切片的分析显示，在从前期到后期的纺锤体极有大量中心粒周围物质。在纵切面上，纺锤体微管呈现为规则的直线状结构。我们假设γ - 微管蛋白位于微管靠近极的末端，和/或在微管的整个长度上与纺锤体微管相关联。为了区分这些可能性，对实验室物种粉斑螟（螟蛾科）的睾丸进行冷处理，然后用抗β - 微管蛋白和γ - 微管蛋白的抗体进行双重标记。预期这种处理会使微管解聚。由含γ - 微管蛋白的物质以末端方式成核的星体微管确实解聚了。相反，含γ - 微管蛋白的纺锤体微管仍然存在。因此，可以想象在鳞翅目物种的雄性减数分裂中，γ - 微管蛋白在微管的整个长度上与微管相关联。这种关联使微管稳定，防止冷诱导的解聚，这似乎是合理的。
