STUDIES ON THE MECHANISM OF PHOSPHATE ACCUMULATION BY SEA URCHIN EMBRYOS

¹ Department of Zoology and the Friday Harbor Laboratories, University of Washington, Seattle 5, Washington

1. The accumulation of phosphate by the eggs and embryos of the sea urchin, Strongylocentrotus purpuratus (Stimpson) was analyzed by determining the action of various metabolic effectors on the uptake of P³² from sea water flowing at a constant rate over the eggs in a special perfusion chamber.

2. The rate of uptake of P³² by unfertilized eggs is nearly zero. The rate for the first 7 to 30 minutes after fertilization (lag phase) is also nearly zero, but increases rapidly during the next 20 to 30 minutes (augmentation phase), and becomes maximal 22 to 60 minutes after fertilization (accumulation phase) at a level many times that of the unfertilized eggs.

3. Artificial parthenogenesis, by either the single or double treatment, results in the same pattern and magnitude of uptake as does fertilization, even in the absence of cleavage.

4. Phosphate accumulation is markedly inhibited by l0^-4 M 2,4-dinitrophenol if this agent is added during the lag phase, moderately inhibited if added during the augmentation phase, but is unaffected if added during the accumulation phase.

5. Addition of 10^-4 M ATP simultaneously with dinitrophenol early in the lag phase does not alleviate the inhibition caused by the latter.

6. ATP neither penetrates into cleaving eggs nor is hydrolyzed by an ATPase at their surface.

7. 10^-4 M arsenate markedly inhibits P³² uptake at all times after fertilization.

8. Arsenate does not materially retard early development of these sea urchin eggs indicating that it does not penetrate into them.

9. P³² uptake has a temperature coefficient of 2 to 2.3 during the accumulation phase.

10. The evidence resulting from the use of these effectors indicates that P³² uptake in sea urchin embryos is enzymatically controlled and that the enzymatic mechanism is located on the cell surface. The period immediately following fertilization is believed to be a time when the uptake mechanism is being established. This process appears to be dependent on phosphate bond energy, the production of which is DNP-sensitive. During the accumulation phase it is suggested that the energy requirements for the operation and maintenance of this mechanism are quantitatively much smaller, or are satisfied by phosphate bond energy the production of which is DNP-insensitive. Possible reasons for this difference in sensitivity are discussed.