| Summary: | 碩士 === 國立臺灣海洋大學 === 生物科技研究所 === 102 === Heat shock protein 60 (Hsp60), a member of the mitochondrial chaperonin family, is highly conserved cellular stress proteins present in every organism from bacteria to man. In mammalians, Hsp60 have been shown to be highly expressed in early stages of germline cells in both sexes, suggesting that it might play an important role during gametogenesis. In invertebrate, however, little research thus far has been conducted on Hsp60 in germline cells. To accumulate the knowledge of Hsp60 in reproduction, in the present study, we investigated the expression of Hsp60 in germline cells in the testis and ovary of a scleractinian coral Euphyllia ancora. The full-length of E. ancora hsp60 cDNA was elucidated, and a specific antibody against the Hsp60 was successfully generated. Immunohistochemical analysis of male corals revealed that Hsp60 was strongly expressed in spermatogonia and early spermatocytes, but not in the post meiotic spermatids or spermatozoa. On the other hand, immunohistochemical analysis of female corals revealed that Hsp60 was strongly expressed in oogonia and early oocytes, and weakly expressed in later stage oocytes that exceeded 200 μm in diameter. Immunoreactivity were also detected in the somatic cells in the tentacles and mesenterial filament in both sexes. In addition, western blotting analysis of unfertilized mature eggs proved the presence of Hps60, suggesting that Hsp60 may be a maternal factor. Expression of Hsp60 in germline were also confirmed in other coral species (12 species) assessed by Immunohistochemical analysis. These results demonstrated that Hsp60 is expressed in germline cells in the corals, suggesting that it might play a fundamental role in germ cell development in corals.Furthermore, we hypothesized that Hsp60 expression in germline is evolutionarily conserved throughout metazoans. To verify this hypothesis, with anti-hsp60 antibody we performed immunohistochemical analysis on various metazoans reproductive organs. It was confirmed that Hsp60 were present at a high level in the germline cells of cnidarians (corals, anemone), mollusks (oysters, abalone), arthropod (shrimp), and vertebrates (mouse, frog, fish). These results suggested that Hsp60 might possess an evolutionarily conserved fundamental role in metazoan germline.
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