Mating elicits two major changes in the reproductive behavior of many insect females. The egg-laying rate increases and the readiness to accept males (receptivity) is reduced. These postmating responses last Ϸ1 week in Drosophila melanogaster. Males that do not transfer sperm but transfer seminal fluid during mating induce a short-term response of 1 day. The long-term response of 1 week requires the presence of sperm (sperm effect). Hence, sperm is essential for the long-term persistence of the postmating responses. Three seminal fluid peptides elicit postmating responses: ovulin, sex-peptide (SP), and DUP99B. Using the technique of targeted mutagenesis by homologous recombination, we have produced males with mutant SP genes. Here, we report that males lacking functional SP elicit only a weak short-term response. However, these males do transfer sperm. Thus, (i) SP is the major agent eliciting the short-term and the long-term postmating responses and (ii) sperm is merely the carrier for SP. The second conclusion is supported by the finding that SP binds to sperm. The 36-aa-encoding SP gene is the first small Drosophila gene knocked out with the method of homologous recombination. I n many insect species, the reproductive behavior of females drastically changes after mating (1, 2). Egg laying is increased and receptivity is reduced. These female postmating responses are elicited by seminal fluid and sperm transferred during copulation. In most species, the virgin state is achieved again after some time (2). The egg-laying rate decreases to a virgin level and the female accepts courting males again. The coordination of egg production and egg laying with the presence of sperm in the female genital tract is largely in the interest of both sexes. However, it may be advantageous for a female to mate with more than one male to enhance the genetic diversity of her offspring. Female promiscuity can reduce the partner's fitness through sperm competition. As countermeasures, males have evolved mechanisms that reduce the females' tendency to remate (2-4). In some species, this may even lead to a reduced female lifespan (5). Experimental approaches have yielded insights into the nature of these sexual conflicts (6-11), but the molecular mechanisms involved remain largely unknown.In Drosophila melanogaster, two male peptides induce the postmating responses when injected into virgin females: sexpeptide (SP, ACP70A) and ductus ejaculatorius peptide 99B (DUP99B; 99B stands for the cytological localization of the gene at position 99B; refs. 12 and 13). They are synthesized in the accessory glands and the ductus ejaculatorius, respectively. Both are transferred into the female during mating (3, 13). After entering the female genital tract, the two peptides are believed to be transferred into the hemolymph where they reach their targets (3,(13)(14)(15). This view is supported by ectopic, constitutive expression of SP in the fat body of females, leading to a continuous induction of the postmating responses (16). Although both peptid...

By binding to sperm tails, SP prolongs the PMR. Thus, besides a carrier for genetic information, sperm is also the carrier for SP. Binding to sperm may protect the peptide from degradation by proteases in the hemolymph and, thus, prolong its half-life. Longer sperm tails may transfer more SP and thus increase the reproductive fitness of the male. We suggest that this could explain the excessive length of sperm tails in some Drosophila species.

Seminal fluid elicits a variety of physiological and behavioral changes in insect females. In Drosophila melanogaster females, sex peptide (SP) is the major seminal agent eliciting oviposition and reduction of receptivity. But SP also has many other effects; for example, it stimulates food intake, egg production, ovulation, juvenile hormone production and antimicrobial peptide synthesis. Thus, SP very probably has several receptors. To identify putative targets and signaling cascades, we studied the genome-wide regulation of genes by microarray analysis of RNA isolated from females after mating with wild-type males or males lacking SP, respectively. In addition, we studied the effects of SP on the proteome of females. Sex peptide regulates gene activity differentially in the head and in the abdomen. Genes coding for unspecific antimicrobial peptides are specifically transcribed in the abdomen, e.g. the antimicrobial peptide drosocin in epithelial tissues of the female genital tract (oviduct and calyx). Hence, SP elicits a systemic [Peng J, Zipperlen P & Kubli E (2005) Curr Biol15, 1690-1694] and an epithelial immune response. Ectopic expression of SP in the fat body of transgenic virgin females (with subsequent secretion into the hemolymph) does not elicit drosocin synthesis in the genital tract. Thus, the receptors for the stimulation of the systemic and the epithelial responses by SP are compartmentalized. The hydroxyproline (P*) motif of SP, P*TKFP*IP*SP*NP*, is identified as a novel elicitor of the innate immune response. We suggest that SP acts by chemical mimicry of sugar components of the bacterial cell wall. Thus, SP may induce the immune system via pattern recognition receptors.

Mating elicits two postmating responses in many insect females: the egg laying rate increases and sexual receptivity is reduced. In Drosophila melanogaster, two peptides of the male genital tract, sex-peptide and DUP99B, elicit these postmating responses when injected into virgin females. Here we show that the gene encoding DUP99B is expressed in the male ejaculatory duct and in the cardia of both sexes. The DUP99B that is synthesized in the ejaculatory duct is transferred, during mating, into the female genital tract. Expression of the gene is first seen in a late pupal stage. Although such males synthesize DUP99B in wild-type quantities, they elicit only weak postmating responses in their mating partners. Males lacking the Dup99B gene elicit the two postmating responses to the same extent as wild-type males. These results suggest that both sexpeptide and DUP99B can elicit both responses in vivo. However, sex-peptide seems to play the major role in eliciting the postmating responses, while DUP99B may have specialized for other, as yet unknown, functions.