By combining elements of two Y-autosome translocations with displaced autosomal breakpoints, it is possible to produce zygotes heterozygous for a deficiency for the region between the breakpoints, and also, as a complementary product, zygotes carrying a duplication for precisely the same region. A set of Y-autosome translocations with appropriately positioned breakpoints, therefore, can in principle be used to generate a non-overlapping set of deficiencies and duplications for the entire autosomal complement.—Using this method, we have succeeded in examining segmental aneuploids for 85% of chromosomes 2 and 3 in order to assess the effects of aneuploidy and to determine the number and location of dosage-sensitive loci in the Drosophila genome (Figure 5). Combining our data with previously reported results on the synthesis of Drosophila aneuploids (see Lindsley and Grell 1968), the following generalities emerge.—1. The X chromosome contains no triplo-lethal loci, few or no haplo-lethal loci, at least seven Minute loci, one hyperploid-sensitive locus, and one locus that is both triplo-abnormal and haplo-abnormal. 2. Chromosome 2 contains no triplo-lethal loci, few or no haplo-lethal loci, at least 17 Minute loci, and at least four other haplo-abnormal loci. 3. Chromosome 3 contains one triplo-lethal locus that is also haplo-lethal, few or no other haplo-lethal loci, at least 16 Minute loci, and at least six other haplo-abnormal loci. 4. Chromosome 4 contains no triplo-lethal loci, no haplo-lethal loci, one Minute locus, and no other haplo-abnormal loci.—Thus, the Drosophila genome contains 57 loci, aneuploidy for which leads to a recognizable effect on the organism: one of these is triplo-lethal and haplo-lethal, one is triplo-abnormal and haplo-abnormal, one is hyperploid-sensitive, ten are haplo-abnormal, 41 are Minutes, and three are either haplo-lethals or Minutes. Because of the paucity of aneuploid-lethal loci, it may be concluded that the deleterious effects of aneuploidy are mostly the consequence of the additive effects of genes that are slightly sensitive to abnormal dosage. Moreover, except for the single triplo-lethal locus, the effects of hyperploidy are much less pronounced than those of the corresponding hypoploidy.

Previous work has established that the polyspermy block in Urechis acts at the level of sperm-egg membrane fusion . (J. Exp . Zool. 196:105) . Present results indicate that during the first 5-10 min after insemination the block is mediated by a positive shift in membrane potential (the fertilization potential) elicited by the penetrating sperm, since holding the membrane potential of the unfertilized egg positive by passing current reduces the probability of sperm entry, while progressively reducing the amplitude of the fertilization potential by decreasing external Na' progressively enhances multiple sperm penetrations . Also, a normal fertilization potential is correlated with a polyspermy block even under conditions (pH 7) in which eggs do not develop . We have investigated the mechanism of the electrical polyspermy block by quantifying the relationship between sperm incorporation, membrane potential and ion fluxes . Results indicate that the polyspermy block is mediated by the electrical change per se and not by the associated fluxes of Na', Ca", and H' . KEY WORDS polyspermy prevention membrane fusion regulation membrane potential ion fluxes fertilizationNormal embryonic development requires that a single sperm nucleus unite with the egg nucleus (see reference 1 for a review of strategies for monospermic fertilization) . In the eggs of Urechis, a significant reduction in susceptibility to polyspermic fertilization occurs within seconds after insemination (reference 38, and our results below) . Paul's results (38) and ours below clearly show that this polyspermy block, rather than being absolutely effective as the term implies, operates by reducing the probability of a second sperm entry 426 sufficiently so that, under presumed physiological sperm-egg ratios, most eggs are monospermic. Although fertilized Urechis eggs become significantly more resistant to refertilization within seconds after insemination, they remain indistinguishable from unfertilized eggs by light and electron microscopy through 4 min after insemination (13), except for a rounding-out of the cell shape (the unfertilized egg has a single, large indentation) . At 4 min the surface coat begins to elevate, but there is no massive exocytosis of cortical vesicles, most of which remain intact through first cleavage (13,39) . Though elevated, the surface coat is no barrier to supernumerary sperm, whose acrosomal tubules penetrate to within 100 A of the egg plasma mem-J . CELT brane (40). Furthermore, sperm can fertilize eggs whose surface coats have been experimentally elevated before insemination (using trypsin at pH 7) .' Thus, it is apparent that the Urechis polyspermy block acts by altering the egg plasma membrane to prevent fusion with supernumerary sperm. In this paper, we show that the fertilization potential, a positive shift in egg membrane potential elicited by the fertilizing spermatozoon, mediates the polyspermy block during the first 5-10 min after insemination . Such an electrical polyspermy block was first demonstrated in ...

. Dr. Jaffe is presently at the Marine Biological Laboratory, Woods Hole, Massachusetts 02543.A a S X R A C x Microelectrode and tracer flux studies of the Urechis egg during fertilization have shown: (a) insemination causes a fertilization potential; the membrane potential rises from an initial level of -33 -+ 6 mV to a peak at +51 -+ 6 mV (n = 16), falls to a plateau of about +30 mV, then returns to the original resting potential 9 -+ 1 min (n = 10) later; (b) the fertilization potential results from an increase in Na + permeability, which is amplified during the first 15 s by a Ca ++ action potential; (c) the maximum amplitude of the fertilization potential, excluding the first 15 s, changes by 51 mV for a 10-fold change in external [Na+]; (d) in the 10 min period after insemination, both Na + and Ca ++ influxes increase relative to unfertilized egg values by factors of 17 -+ 7 (n = 6) and 34 -+ 14 (n = 4), respectively; the absolute magnitude of the Na + influx is 16 _+ 6 times larger than that of Ca++; (e) in the absence of sperm these same electrical and ionic events are elicited by trypsin; thus, the ion channels responsible must preexist in the unfertilized egg membrane; 0 c) increased Na + influx under conditions of experimentally induced polyspermy indicates that during normal monospermic fertilization, only a fraction of available Na + channels are opened; we conclude that these channels are sperm-gated; (g) Ca ++ influx at fertilization is primarily via the membrane potential-gated channel, because kinetics are appropriate, and influx depends on potential in solutions of varying [Na+], but is independent of number of sperm incorporations in normal sea water.

A method for the organ culture ofDrosophila testes is described which supports the differentiation of primary spermatocytes through the meiotic divisions to elongating spermatids. Autoradiographic and inhibitor studies reveal no evidence for RNA synthesis by developing spermatids ofDrosophila melanogaster; most, if not all, of the RNA required for the differentiation and elongation of sperm is synthesized earlier in the primary spermatocytes. Primary spermatocytes will differentiate into elongating spermatids in organ culture, despite severe (96-98%) inhibition ofH-uridine incorporation into RNA effected by 50 μg/ml 3'-deoxyadenosine. Protein synthesis in spermatids continues to be active in the presence of 3'-deoxyadenosine, but that in growing spermatocytes is severely inhibited.