ASCIDIAN EGGS BLOCK POLYSPERMY BY 2 INDEPENDENT MECHANISMS - ONE AT THE EGG PLASMA-MEMBRANE, THE OTHER INVOLVING THE FOLLICLE CELLS

Many ascidians live in clumps and usually release sperm before the egg s. Consequently, eggs are often spawned into dense clouds of sperm. Be cause fertilization by more than a single sperm is lethal, ascidians h ave evolved at least two successive blocks to polyspermy: the rapid re lease of a glycosidase that inhibits sperm binding to the vitelline co at (VC) and a subsequent change in membrane potential that prevents su pernumerary sperm-egg fusion. This paper shows that (1) these two bloc ks can be uncoupled by the use of suramin, and (2) most of the glycosi dase appears to be from the follicle cells, which are accessory cells on the outside of the egg VC. Phallusia mammillata eggs initially bind numerous sperm but, after the glycosidase is released, only a few add itional sperm bind. Intact eggs in 20 mu M suramin release glycosidase , but the electrical response is inhibited; sperm swim actively and bi nd to the VC but fail to penetrate. Suramin treatment is completely re versible; intact eggs exhibit the electrical response an average of 11 minutes after the drug is washed out. Sperm must contact the follicle cells before passing through the VC; eggs with the VC removed and fer tilized in the presence of 20 mu M suramin show the electrical respons e 35% of the time, thus VC removal enhances sperm entry. Like the inta ct eggs, 100% of the naked eggs respond electrically to fertilization after the drug is washed out. Follicle cells that are isolated by calc ium magnesium free seawater and then returned to complete seawater rel ease N-acetylglucosaminidase activity in response to sperm. Thus, thes e eggs have two blocks to polyspermy that operate in sequence: an earl y first block resulting from enzymatic modification of the VC by N-ace tylglucosaminidase released primarily from follicle cells and a second electrical block operating at the egg plasma membrane level and requi ring sperm-egg fusion. (C) 1997 Wiley-Liss, Inc.