Studies of the mechanism of the electrical polyspermy block using voltage clamp during cross-species fertilization.

Jaffe, Laurinda A.; Gould-Somero, Meredith; Holland, Linda Z.

doi:10.1083/jcb.92.3.616

Cited by 54 publications

(19 citation statements)

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“…In some animals (sea urchins and frogs being the best studied), the block at the membrane is accomplished by a fusion-induced depolarization of the egg plasma membrane that through unknown mechanisms disallows penetration of the egg by additional sperm (Jaffe et al, 1982). In mammals, the membrane block to polyspermy is slower (30-60 minutes) and requires sperm membrane fusion with the egg, but otherwise is uncharacterized (Wortzman-Show et al, 2007).…”

Section: Introductionmentioning

confidence: 99%

Membrane fusion triggers rapid degradation of two gamete-specific, fusion-essential proteins in a membrane block to polygamy inChlamydomonas

Liu

Misamore

Snell³

2010

Development

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SUMMARYThe plasma membranes of gametes are specialized for fusion, yet, once fusion occurs, in many organisms the new zygote becomes incapable of further membrane fusion reactions. The molecular mechanisms that underlie this loss of fusion capacity (block to polygamy) remain unknown. During fertilization in the green alga Chlamydomonas, the plus gamete-specific membrane protein FUS1 is required for adhesion between the apically localized sites on the plasma membranes of plus and minus gametes that are specialized for fusion, and the minus-specific membrane protein HAP2 is essential for completion of the membrane fusion reaction. HAP2 (GCS1) family members are also required for fertilization in Arabidopsis, and for the membrane fusion reaction in the malaria organism Plasmodium berghei. Here, we tested whether Chlamydomonas gamete fusion triggers alterations in FUS1 and HAP2 and renders the plasma membranes of the cells incapable of subsequent fusion. We find that, even though the fusogenic sites support multi-cell adhesions, triploid zygotes are rare, indicating a fusion-triggered block to the membrane fusion reaction. Consistent with the extinction of fusogenic capacity, both FUS1 and HAP2 are degraded upon fusion. The rapid, fusion-triggered cleavage of HAP2 in zygotes is distinct from degradation occurring during constitutive turnover in gametes. Thus, gamete fusion triggers specific degradation of fusion-essential proteins and renders the zygote incapable of fusion. Our results provide the first molecular explanation for a membrane block to polygamy in any organism.

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Section: Introductionmentioning

confidence: 99%

Membrane fusion triggers rapid degradation of two gamete-specific, fusion-essential proteins in a membrane block to polygamy inChlamydomonas

Liu

Misamore

Snell³

2010

Development

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“…The fertilization potential has a duration of 10-20 min and provides a fast, transient block to polyspermy (Cross & Elinson, 1980;Grey, Bastiani, Webb & Schertel, 1982;Jaffe, Cross & Picheral, 1983). The positive shift is caused by an increase in Cl-permeability (Maeno, 1959;Cross, 1981) and the presence of a Cl-gradient (44 mm inside, Jaffe & Schlichter, 1985, and a few millimolar outside).…”

Section: Introductionmentioning

confidence: 99%

Propagating potassium and chloride conductances during activation and fertilization of the egg of the frog, Rana pipiens.

Jaffe¹,

Kado²,

L³

1985

The Journal of Physiology

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SUMMARY1. Fertilization or artificial activation of the frog egg (Rana pipiens) elicits a positive-going shift in membrane potential which results from an opening of Cl-and K+ channels in the egg membrane. We examined the spatial localization of the currents produced by the opening of these channels, using large patch electrodes (tip diameters 3-10 ,um). We also used small patch electrodes (tip diameters about 1 /Im) to study the single K+ channel currents.2. After activation, with the patch electrode at any position on the egg surface, we observed a transient current, with a main peak lasting several seconds. This activation current occurred after a variable delay of 0-3 min following the rise of the activation potential. With 10 % Ringer solution in the bath and pipette, the current was usually outward, although it sometimes had an inward component.3. With one patch electrode on the animal surface of the egg and another patch electrode on the vegetal surface, we observed that the activation current propagated over the egg surface. In experiments where the egg was activated by applying a hyperpolarizing pulse, the response in the animal half preceded that in the vegetal half by an average of about 1 min. The amplitude of the peak outward current was similar for animal and vegetal recordings (1-2 mA/cm2).4. Tetraethylammonium (11 mM) in the patch pipette blocked most ofthe outward component of the activation current and revealed an underlying inward component.5. The inward component of the activation current was carried by Cl-, since it could be reversed by raising the Cl-concentration in the pipette.6. The Cl-component of the activation current propagated over the egg surface, with timing similar to that of the total current. 7. The average amplitude of the peak Cl-current was six or more times larger at the animal than the vegetal surface.8. Fertilization caused a current to propagate from the animal to the vegetal surface, like the current observed during activation.9. With a small patch electrode, single channel currents of the K+ component of the activation current could be seen. The probability that the channels were open increased at more positive potentials. The single channel conductance was estimated 8-2 228 L. A. JAFFE, R. T. KADO AND L. MUNCY to be 25 pS, and the reversal potential to be -150 mV. Single Cl-channel currents have not yet been seen.10. Activation or fertilization of the frog egg resulted in a wave-like opening of Cl-and K+ channels, which spread from the animal to the vegetal half of the egg. The Cl-conductance was larger on the animal half.

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“…Since the voltage required to block fertilization depends on the species of sperm, the best model for the electrical block is a fusion or transfer process in which some component of the sperm is physically inserted into the egg membrane so that it is sufficiently across the resistance of the egg membrane to experience the membrane potential (8,9). We have used this model in discussing our results on overcoming the electrical block.…”

Section: Discussionmentioning

confidence: 99%

“…Although there is no direct evidence on how the positive inside values of egg membrane potential block the completion of fusion of the sperm with the egg and the activation of the egg, there is suggestive evidence that the voltage-sensitive component is a property of the sperm and not the egg membrane (8,9). This implies that the electrical block is directly interfering with the fusion process itself, because the voltage-sensitive component of the sperm can be influenced by the egg membrane potential only when it is inserted at least partially across the egg membrane resistance.…”

mentioning

confidence: 99%

“…However, the inside-positive values of membrane potential that result from fertilization by the first sperm can serve as the basis for the fast block to polyspermy in the eggs of sea urchins (1), the marine worm Urechis (6), and the frog Xenopus (7). The electrical block can also be artificially imposed by the application of current through an intracellular electrode so that the inside-positive membrane potentials prevent any sperm from completing the fusion and activation process (1).Although there is no direct evidence on how the positive inside values of egg membrane potential block the completion of fusion of the sperm with the egg and the activation of the egg, there is suggestive evidence that the voltage-sensitive component is a property of the sperm and not the egg membrane (8,9). This implies that the electrical block is directly interfering with the fusion process itself, because the voltage-sensitive component of the sperm can be influenced by the egg membrane potential only when it is inserted at least partially across the egg membrane resistance.…”

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confidence: 99%

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Time and voltage windows for reversing the electrical block to fertilization.

Shen¹,

Steinhardt²

1984

Proc. Natl. Acad. Sci. U.S.A.

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The electrical block to fertilization of sea urchin eggs can be overcome by very brief periods of inside-negative egg membrane potential. Lytechinus Pictus eggs whose membrane potentials have been clamped at + 15 mV cannot be fertilized. If the membrane potential is repolarized to insidenegative voltages for a brief interval, the egg can be successfully fertilized. By varying the duration and voltage of these brief periods of inside negativity, we have uncovered three general properties of the electrically sensitive step in fertilization.First, a membrane-potential step that becomes rate limiting at inside-positive voltages can be initiated within a few milliseconds of inside negativity (30-60 msec at -60 mV). Second, at the time that the electrically sensitive step is being completed, there are other potential-independent steps with probably slower time constants because the duration of negativity was more effective applied as paired pulses rather than a single long pulse. Third, the permissive state is more quickly established by inside negativity than the nonpermissive state is established by inside positivity because the interval between paired pulses could be a few times longer than the effective single pulse in duration. In these voltage-clamped eggs the intervals from the successful completion of the electrically sensitive step to the next identifiable signs of activation were on the order of several seconds and highly variable.At fertilization in a sea urchin egg, the first successful sperm triggers rapid permeability increases to calcium and sodium that reverse the polarity of the egg membrane potential to inside-positive values (1-4). This rapid depolarization to inside-positive membrane potentials prevents fertilization by secondary sperm (1). This is called the electrical block. These permeability changes do not appear to play a direct role in the activation of development (5). However, the inside-positive values of membrane potential that result from fertilization by the first sperm can serve as the basis for the fast block to polyspermy in the eggs of sea urchins (1), the marine worm Urechis (6), and the frog Xenopus (7). The electrical block can also be artificially imposed by the application of current through an intracellular electrode so that the inside-positive membrane potentials prevent any sperm from completing the fusion and activation process (1).Although there is no direct evidence on how the positive inside values of egg membrane potential block the completion of fusion of the sperm with the egg and the activation of the egg, there is suggestive evidence that the voltage-sensitive component is a property of the sperm and not the egg membrane (8,9). This implies that the electrical block is directly interfering with the fusion process itself, because the voltage-sensitive component of the sperm can be influenced by the egg membrane potential only when it is inserted at least partially across the egg membrane resistance. In the experiments described here, we have attempted to furt...

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Studies of the mechanism of the electrical polyspermy block using voltage clamp during cross-species fertilization.

Cited by 54 publications

References 21 publications

Membrane fusion triggers rapid degradation of two gamete-specific, fusion-essential proteins in a membrane block to polygamy inChlamydomonas

Membrane fusion triggers rapid degradation of two gamete-specific, fusion-essential proteins in a membrane block to polygamy inChlamydomonas

Propagating potassium and chloride conductances during activation and fertilization of the egg of the frog, Rana pipiens.

Time and voltage windows for reversing the electrical block to fertilization.

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