Molecular Mechanism Of Two Cytoskeleton And Related Motor Protein During Spermatogenesis Of Macrobrachium Nipponense

Live-cell imaging, immunofluorescence, and electron microscopy have paved the way for the advances in understanding spermatogenesis of Decapoda, Crustacea from the ultrastructural level. Spermatogenesis is a complex differentiation process, heavily dependent on the cytoskeletal organization and the motor proteins. Macrobrachium nipponense is representative of the Caridean shrimp, the sperm of which is among the typical aflagellate ones and has a highly specialized morphology, resembling a thumb tack. The mature sperm consists of a shallow-dish-like main body and a spike. There is no specific research concerning the forming of this particular kind of acrosome, and the molecular mechanism of cytoskeleton and its motor protein during nucleus variation of Macrobrachium nipponense has not been studied. To probe how this molecular mechanism helps shape the aflagellate sperm is our aim.We used frozen sections of testis and spermaduct et al. of wild Macrobrachium nipponense, at different developmental stages, which were studied by means of immunofluorescence to find out the distribution of actin, tubulin, and microtubule-related motor protein KIFC1. In addition to this, we also analyzed the protein expression of KIFC1 in different tissues by western blotting, and this was performance for the first time in Macrobrachium nipponense.The western blotting results showed that KIFC1 expressed in all tissues we analyzed, and had no tissue-differential expression. The results of immunofluorescence indicated that microtubule and KIFC1 may play important roles in forming the lamellar complex, and they worked together to help transport the vesicle and protein complex during spermatocyte stage and early spermatid stage. While at the stage of late spermatid and sperm, the distribution of KIFC1 had gradually shifted from cytoplasm to nucleoplasm, showing that KIFC1 may well be to associate with the differentiation of chromatin, which would develop into two types of nucleus in the mature sperm, i.e., vesicular nucleus and filament nucleus. At the same time, a microtubule-based structure named "acroframosome" appears at the acrosome site, having two bright immunofluorescence spots in its center. The two spots of highest brightness were supposed to be MTOC, where the microtubule concentrated. The acroframosome may function like manchette in mammalian germ cell during acrosome formation. The actin was confirmed to exist in testis of Macrobrachium nipponense according to the result of immunofluorescence, and could reasonably play some part during spermatogenesis. The actins were arranged in network, surrounding the nucleus. But still, the underlying mechanism needs further exploring.