Department of Anatomy and Developmental Biology, Graduate School of Medicine, Chiba University, Chiba 260-8670, Japan.
Reproduction. 2010 Mar;139(3):533-44. doi: 10.1530/REP-09-0434. Epub 2009 Dec 23.
It is important to establish a reliable and progressive model of the acrosome reaction. Here, we present a progression model of the acrosome reaction centering around the acrosomal membrane-anchored protein equatorin (MN9), comparing the staining pattern traced by MN9 antibody immunofluorescence with that traced by Arachis hypogaea agglutinin (PNA)-FITC. Prior to the acrosome reaction, equatorin was present in both the anterior acrosome and the equatorial segment. Since sperm on zona pellucida showed various staining patterns, MN9-immunostaining patterns were classified into four stages: initial, early, advanced, and final. As the acrosome reaction progressed from the initial to the early stage, equatorin spread from the peripheral region of the anterior acrosome toward the center of the equatorial segment, gradually over the entire region of the equatorial segment during the advanced stage, and finally uniformly at the equatorial segment at the final stage. In contrast, the PNA-FITC signals spread more quickly from the peripheral region of the acrosome toward the entire equatorial segment, while decreasing in staining intensity, and finally became weak at the final stage. MN9-immunogold electron microscopy showed equatorin on the hybrid vesicles surrounded by amorphous substances at advanced stage of acrosome reaction. Equatorin decreased in molecular mass from 40-60 to 35 kDa, and the signal intensity of 35 kDa equatorin increased as the acrosome reaction progressed. Thus, the established equatorin-based progression model will be useful for analyzing not only the behavior of equatorin but also of other molecules of interest involved in the acrosome reaction.
建立可靠且渐进的顶体反应模型非常重要。在这里，我们提出了一个以顶体膜锚定蛋白赤道蛋白（MN9）为中心的顶体反应进展模型，比较了 MN9 抗体免疫荧光追踪的模式与花生凝集素（PNA）-FITC 追踪的模式。在顶体反应之前，赤道蛋白存在于顶体的前区和赤道段。由于透明带中的精子显示出各种染色模式，因此 MN9 免疫染色模式被分为四个阶段：初始、早期、晚期和终期。随着顶体反应从初始阶段向早期阶段进展，赤道蛋白从前区的周边区域向赤道段的中心扩散，在晚期阶段逐渐覆盖赤道段的整个区域，最后在终期阶段均匀地分布在赤道段。相比之下，PNA-FITC 信号从顶体的周边区域更快地扩散到整个赤道段，同时染色强度逐渐降低，最后在终期阶段变得很弱。MN9-免疫胶体金电子显微镜显示，在顶体反应的晚期，赤道蛋白位于被无定形物质包围的杂交小泡上。赤道蛋白的分子量从 40-60 减少到 35 kDa，随着顶体反应的进展，35 kDa 赤道蛋白的信号强度增加。因此，建立的基于赤道蛋白的进展模型不仅对赤道蛋白的行为，而且对其他参与顶体反应的感兴趣分子的行为分析都将非常有用。
