A Novel Function for the Hox Gene Abd-B in the Male Accessory Gland Regulates the Long-Term Female Post-Mating Response in Drosophila

Abstract

In insects, products of the male reproductive tract are essential for initiating and maintaining the female post-mating response (PMR). The PMR includes changes in egg laying, receptivity to courting males, and sperm storage. In Drosophila, previous studies have determined that the main cells of the male accessory gland produce some of the products required for these processes. However, nothing was known about the contribution of the gland's other secretory cell type, the secondary cells. In the course of investigating the late functions of the homeotic gene, Abdominal-B (Abd-B), we discovered that Abd-B is specifically expressed in the secondary cells of the Drosophila male accessory gland. Using an Abd-B BAC reporter coupled with a collection of genetic deletions, we discovered an enhancer from the iab-6 regulatory domain that is responsible for Abd-B expression in these cells and that apparently works independently from the segmentally regulated chromatin domains of the bithorax complex. Removal of this enhancer results in visible morphological defects in the secondary cells. We determined that mates of iab-6 mutant males show defects in long-term egg laying and suppression of receptivity, and that products of the secondary cells are influential during sperm competition. Many of these phenotypes seem to be caused by a defect in the storage and gradual release of sex peptide in female mates of iab-6 mutant males. We also found that Abd-B expression in the secondary cells contributes to glycosylation of at least three accessory gland proteins: ovulin (Acp26Aa), CG1656, and CG1652. Our results demonstrate that long-term post-mating changes observed in mated females are not solely induced by main cell secretions, as previously believed, but that secondary cells also play an important role in male fertility by extending the female PMR. Overall, these discoveries provide new insights into how these two cell types cooperate to produce and maintain a robust female PMR.

Ciona intestinalis NADH dehydrogenase NDX confers stress-resistance and extended lifespan on Drosophila

“…However, AOX lines created subsequently by targeted integration [A. Andjelkovic, unpublished data] retain full activity. One of the integration sites used in the present study (VK00001 site on chromosome 2) may be subject to position effects [40], although this site has recently been used for functionally successful insertion of a specific GAL4 driver [47]. Moreover, integration on chromosome 3 produced a similar amount of functionality (data not shown).…”

Neprilysins: An Evolutionarily Conserved Family of Metalloproteases That Play Important Roles in Reproduction in Drosophila

“…Fertility/fecundity assays and sperm competition Fly stocks and media: All flies were raised at room temperature (23°6 1°) in glass bottles on standard yeastglucose media (Gligorov et al 2013). Females were aged 3-5 days from eclosion in groups of 5-12 in glass vials with added yeast.…”

“…Different tissues were incubated in a volume of 150 ml proteinase K mixture: inverted third-instar larvae, 15 mg/ml proteinase K for 2 min at 37°; adult abdomen, 15 mg/ml proteinase K for 3 min at 37°; adult thorax, 10 mg/ml proteinase K for 2 min at 37°; adult brain, 10 mg/ml proteinase K for 2 min at 37°; and whole-mount embryos, 40 mg/ml proteinase K for 3 min at RT. Prehybridization, hybridization, and detection were as described in Clements et al (2008).Fertility/fecundity assays and sperm competition Fly stocks and media: All flies were raised at room temperature (23°6 1°) in glass bottles on standard yeastglucose media (Gligorov et al 2013). Females were aged 3-5 days from eclosion in groups of 5-12 in glass vials with added yeast.…”

“…The male was removed upon dismounting. Assays for the effects of the Neps on female fertility were performed the same way using 3-to 5-day-old Canton-S males as mates for either RNAi (or mutant) or control females.After mating, individual females were housed on yeast glucose media for 24 hr after which each female was transferred to a fresh vial, and the eggs laid in the previous vial were counted as described in Gligorov et al (2013) except that Nep1 and Nep4 RNAi and control females were housed at 30°for the entire experiment to ensure adequate suppression of GAL80. The progeny present in each vial were counted after eclosion.…”

A Drosophila Protease Cascade Member, Seminal Metalloprotease-1, Is Activated Stepwise by Male Factors and Requires Female Factors for Full Activity

“…Additionally, Sfps are necessary for sperm function, although they do not form part of the sperm itself. In D. melanogaster, ablation of the major Sfp-producing tissue, the male accessory gland (AG), results in the complete loss of male fertility (Kalb et al 1993;Gligorov et al 2013), unless Sfps are provided by a second male (Xue and Noll 2000). Over 200 Sfps are transferred (or inferred to be transferred) to females during mating in D. melanogaster (Ravi Ram and Wolfner 2007;Findlay et al 2008Findlay et al , 2009Takemori and Yamamoto 2009;Yamamoto and Takemori 2010).…”