Mitochondria-Specific Monoclonal Antibodies in Eggs and Embryos of the Ascidian Halocynthia roretzi
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Ascidian embryos have become an important model for embryological studies, offering a simple example for mechanisms of cytoplasmic components segregation. It is a well-known example that the asymmetric segregation of mito-chondria into muscle lineage cells occurs during ascidian embryogenesis. However, it is still unclear which signaling pathway is involved in this process. To obtain molecular markers for studying mechanisms involved in the asymmetric distribution of mitochondria, we have produced monoclonal antibodies, Mito-1, Mito-2 and Mito-3, that specifically recognize mitochondria-rich cytoplasm in cells of the ascidian Halocynthia roretzi embryos. These antibodies stained cytoplasm like reticular structure in epidermis cells, except for nuclei, at the early tailbud stage. Similar pattern was observed in vital staining of mitochondria with DiOC2, a fluorescent probe of mitochondria. Immunostaining with these antibodies showed that mitochondria are evenly distributed in the animal hemisphere blastomeres at cleavage stages, whereas not in the vegetal hemisphere blastomeres. Mi-tochondria were transferred to the presumptive muscle and nerve cord lineage cells of the marginal zone in the vegetal hemi-sphere more than to the presumptive mesenchyme, notochord and endoderm lineage of the central zone. Therefore, it is sug-gested that these antibodies will be useful markers for studying mechanisms involved in the polarized distribution of mito-chondria during ascidian embryogenesis.
Abstract
Ascidian embryos have become an important model for embryological studies, offering a simple example for mechanisms of cytoplasmic components segregation. It is a well-known example that the asymmetric segregation of mito-chondria into muscle lineage cells occurs during ascidian embryogenesis. However, it is still unclear which signaling pathway is involved in this process. To obtain molecular markers for studying mechanisms involved in the asymmetric distribution of mitochondria, we have produced monoclonal antibodies, Mito-1, Mito-2 and Mito-3, that specifically recognize mitochondria-rich cytoplasm in cells of the ascidian Halocynthia roretzi embryos. These antibodies stained cytoplasm like reticular structure in epidermis cells, except for nuclei, at the early tailbud stage. Similar pattern was observed in vital staining of mitochondria with DiOC2, a fluorescent probe of mitochondria. Immunostaining with these antibodies showed that mitochondria are evenly distributed in the animal hemisphere blastomeres at cleavage stages, whereas not in the vegetal hemisphere blastomeres. Mi-tochondria were transferred to the presumptive muscle and nerve cord lineage cells of the marginal zone in the vegetal hemi-sphere more than to the presumptive mesenchyme, notochord and endoderm lineage of the central zone. Therefore, it is sug-gested that these antibodies will be useful markers for studying mechanisms involved in the polarized distribution of mito-chondria during ascidian embryogenesis.
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