Sex peptide is required for the efficient release of stored sperm in mated Drosophila females

Abstract

The Drosophila seminal fluid protein (SFP) sex peptide (SP) elicits numerous post-mating responses, including increased egg laying and decreased sexual receptivity, in the mated female. Unlike other SFPs, which are detectable in mated females for only a few hours post mating, SP is maintained-and its effects are sustained-for several days. The persistence of SP in the mated female's reproductive tract is thought to be a consequence of its binding to, and gradual release from, sperm in storage, which maintains SP's ability to act within the female reproductive tract. Recent studies have shown that several other SFPs, acting in a network, are needed for SP's localization to sperm and are necessary for the efficient release of sperm from storage. This result suggested an additional new role for SP modulating the release of sperm from storage. We tested for this possibility by examining sperm storage parameters in mated females that did not receive SP. We found that while sperm accumulation into storage was unaffected, sperm depletion from storage sites was significantly decreased (or impaired) in the absence of SP. Mates of males expressing a modified SP that is unable to be released from sperm showed a similar phenotype, indicating that release of sperm-bound SP is a necessary component of normal sperm depletion. Additionally, SP null males were more successful in a sperm competitive environment when they were first to mate, which is likely a consequence of higher retention of their sperm due to defective sperm release. Our findings illustrate a direct role for SP in the release of sperm from storage.

Figure 1.—

Progression of different conformational changes in the uteri of females mated to SP null or control males. Mates of SP null and control males had a similar uterine stage distribution at 35 min ASM. SP has no significant effects on the distribution of uterine conformation stages (N_cont = 24, N_ΔSP = 22, P > 0.86).

Figure 2.—

Females mated to SP null males retain sperm long term. Mean (+1 SE) number of sperm stored in the seminal receptacles (SR) and spermathecae (Spp) of females mated to SP null (shaded bars) and control (solid bars) males at 2 hr, 4 days, and 10 days ASM. Asterisks indicate a statistically significant difference between female groups. We did not detect a significant difference in the number of sperm stored by mates of SP null males compared to their controls in the seminal receptacles or spermathecae at 2 hr ASM (SR: t-test, t = −1.41, N_cont = 18, N_ΔSP = 13, P = 0.17; Spp: t = −1.32, N_cont = 23, N_ΔSP = 22, P = 0.20), but we observed a significant difference in the number of sperm stored in seminal receptacles and spermathecae at 4 days ASM (SR: t = −12.39, N_cont = 47, N_ΔSP = 47, P < 0.0005; Spp: t = −3.63, N_cont = 37, N_ΔSP = 36, P = 0.001) and in the SR, but not in the Spp at 10 days ASM (SR: t = −4.26, N_cont = 29, N_ΔSP = 34, P <0.0005; Spp: t = 1.11, N_cont = 29, N_ΔSP = 34, P = 0. 274).

Figure 3.—

Females mated to males whose SP is unable to be cleaved from sperm retain sperm longer in the seminal receptacles. The number of sperm was counted in the seminal receptacles of females mated to males expressing a modified SP [see text and Peng et al. (2005a,b)] or their controls at 4 days and 10 days ASM. Asterisk indicates a statistically significant difference between groups. Total sperm stored by mates of SP-TG^QQ mutant males was comparable to controls at 4 days ASM (SR: t = −1.54, N_cont = 17, N_SP-TG^QQ = 19, P = 0.065), but we observed a significant difference in the number of sperm stored in the seminal receptacle at 10 days ASM (SR: Wilcoxon: Z = −2.68, N_cont = 10, N_SP-TG^QQ = 10, P < 0.003).

Figure 4.—

First-mating SP null males sire a higher proportion of progeny than do controls (Wilcoxon: Z = −5.2, N_cont = 28, N_ΔSP = 31, P < 0.0001). Asterisk indicates a statistically significant difference between groups.