Oko R, Maravei D
Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec, Canada.
Biol Reprod. 1994 May;50(5):1000-14. doi: 10.1095/biolreprod50.5.1000.
The perinuclear theca is a cytoskeletal structure that covers the nucleus of mammalian spermatozoa except for a narrow zone around the insertion of the tail. It shows two distinct regions, a subacrosomal layer or perforatorium and, continuing caudally beyond the acrosomic system, the postacrosomal sheath. The main objective of this study was to analyze the protein composition of the bull perinuclear theca and the distribution of some of its proteins. The perinuclear theca was extracted from demembranated and acrosome-depleted sperm heads by alkaline treatment. Gel electrophoresis revealed that the extract was composed of over 20 polypeptide bands, of which the 15.5-, 25-, 28-, 32-, 36-, and 60-kDa bands were most prominent. Polyclonal antibodies prepared against these major polypeptides were screened on Western blots of the theca extracts and were immunolocalized on sections of spermatozoa prepared for electron microscopy. Antibodies against the 15.5- and 60-kDa polypeptides reacted monospecifically with their respective bands on Western blots, and immunogold labeled the perforatorium and the entire perinuclear theca, respectively. Antibodies against the 25-, 28-, 32-, and 36-kDa polypeptides showed distinctive patterns of cross-reactivity with other polypeptides on Western blots, but immunogold labeled exclusively the entire perinuclear theca. In conclusion, this study provides evidence that the protein composition of the bull perinuclear theca is complex: at least six major polypeptides make up this cytoskeletal element. As to their distribution, the 15.5-kDa polypeptide is localized exclusively to the subacrosomal layer, and the other five major polypeptides analyzed in this study appear to be shared between the subacrosomal and post-acrosomal regions of the perinuclear theca.
核周鞘是一种细胞骨架结构，它覆盖着哺乳动物精子的细胞核，但尾部插入处周围有一个狭窄区域除外。它有两个不同的区域，一个顶体下层或穿孔器，以及在顶体系统尾侧延伸的顶体后鞘。本研究的主要目的是分析公牛核周鞘的蛋白质组成及其一些蛋白质的分布。通过碱性处理从去膜和顶体缺失的精子头部提取核周鞘。凝胶电泳显示提取物由20多条多肽带组成，其中15.5 kDa、25 kDa、28 kDa、32 kDa、36 kDa和60 kDa的条带最为明显。针对这些主要多肽制备的多克隆抗体在核周鞘提取物的蛋白质印迹上进行筛选，并在制备用于电子显微镜的精子切片上进行免疫定位。针对15.5 kDa和60 kDa多肽的抗体在蛋白质印迹上与其各自的条带发生单特异性反应，免疫金分别标记穿孔器和整个核周鞘。针对25 kDa、28 kDa、32 kDa和36 kDa多肽的抗体在蛋白质印迹上与其他多肽显示出独特的交叉反应模式，但免疫金仅标记整个核周鞘。总之，本研究提供了证据表明公牛核周鞘的蛋白质组成复杂：至少六种主要多肽构成了这种细胞骨架成分。至于它们的分布，15.5 kDa多肽仅定位于顶体下层，本研究分析的其他五种主要多肽似乎在核周鞘的顶体下和顶体后区域共享。
