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Quantitative changes in yolk protein and other components in the ovary and testis of the sea urchin Pseudocentrotus depressus

T. Unuma^1,*, T. Yamamoto², T. Akiyama², M. Shiraishi² and H. Ohta¹

¹ National Research Institute of Aquaculture, Nansei, Mie 516-0193, Japan
² Inland Station, National Research Institute of Aquaculture, Tamaki, Mie 519-0423, Japan

View larger version (138K): [in a new window]   Fig. 1. Representative morphology in the ovary (A—D) and testis (E—H) of P. depressus at different stages. Paraffin-embedded sections were stained with Hematoxylin and Eosin. At stage 1 (A,E), the gonadal lumina are filled with nutritive phagocytes. At stage 2 (B,F), many developing oocytes or clusters of spermatogonia are present at the periphery. At stage 3 (C,G), nutritive phagocytes are replaced with ripe ova or spermatozoa in the center of the lumina. At stage 4 (D,H; fully mature gonad), the lumina are filled with ripe ova or spermatozoa. NP, nutritive phagocyte; OC, oocyte; OV, ripe ovum; SG, spermatogonium; SC, spermatocyte; SZ, spermatozoon. Scale bar, 100 µm.

View larger version (20K): [in a new window]   Fig. 2. Standard curves of sandwich ELISA for serial twofold dilution of the purified major yolk protein (MYP) and the gonadal extract in P. depressus.

View larger version (17K): [in a new window]   Fig. 3. Changes in the gonad indices in male and female P. depressus at different stages during gametogenesis. Values are means ± S.E.M. obtained from 6 to 36 individuals.

View larger version (24K): [in a new window]   Fig. 4. Quantitative changes in the content of major yolk protein (MYP) (A), water (B), protein (C), lipid (D), polysaccharide (E) and nucleic acids in ovary (F) and testis (G) in the gonad of P. depressus at different stages during gametogenesis. Values are means ± S.E.M. obtained from 3-6 individuals, except for the testes at stage 2 for protein (2 individuals). In some graphs, the S.E.M. values are smaller than the size of the symbols.

View larger version (31K): [in a new window]   Fig. 5. Changes in the biochemical composition of the gonad of P. depressus during gametogenesis. To standardize the change in the gonadal size, the level of each component shown in Fig. 4 was multiplied by the average gonad index at each stage shown in Fig. 3. Thus the values represent the amount of each component of the gonad in animals at a body mass of 100 g. (A) Ovary, (B) testis. Protein includes major yolk protein (MYP), denoted by diagonal stripes, and other proteins.

View larger version (56K): [in a new window]   Fig. 6. Gonadal homogenates, egg homogenate (E), and the purified major yolk protein (P) in P. depressus, subjected to (A) SDS-polyacrylamide gradient (5-20%) gel electrophoresis, stained with Coomassie Brilliant Blue (CBB), and (B) western blot analysis, immunostained with anti-MYP. Samples containing 10 µg of protein were applied to each lane. Numbers indicate the gonad stages. Molecular mass values on the left are indicated by the migration positions of marker proteins. In the western blots, higher molecular mass bands above the main bands are probably aggregations of MYP. Lower molecular mass bands below the main bands are cleavage products of MYP (e.g. Scott and Lennarz, 1989).

View larger version (33K): [in a new window]   Fig. 7. Hypothetical diagram of the nutritional role of major yolk protein (MYP) in sea urchin reproduction. Circles, proteins; hexagons, other molecules such as nucleic acids, lower molecular mass substances, etc. Broken lines with arrows indicate the loss caused by metabolism as an energy source.