Abstract
Across diverse taxa, seminal fluid proteins (Sfps) transferred at mating affect the reproductive success of both sexes. Such reproductive proteins often evolve under positive selection between species; because of this rapid divergence, Sfps are hypothesized to play a role in speciation by contributing to reproductive isolation between populations. In Drosophila, individual Sfps have been characterized and are known to alter male sperm competitive ability and female post-mating behavior, but a proteomic-scale view of the transferred Sfps has been missing. Here we describe a novel proteomic method that uses whole-organism isotopic labeling to detect transferred Sfps in mated female D. melanogaster. We identified 63 proteins, which were previously unknown to function in reproduction, and confirmed the transfer of dozens of predicted Sfps. Relative quantification of protein abundance revealed that several of these novel Sfps are abundant in seminal fluid. Positive selection and tandem gene duplication are the prevailing forces of Sfp evolution, and comparative proteomics with additional species revealed lineage-specific changes in seminal fluid content. We also report a proteomic-based gene discovery method that uncovered 19 previously unannotated genes in D. melanogaster. Our results demonstrate an experimental method to identify transferred proteins in any system that is amenable to isotopic labeling, and they underscore the power of combining proteomic and evolutionary analyses to shed light on the complex process of Drosophila reproduction.
Highlights
In addition to sperm, males of many internally fertilizing species transfer seminal fluid proteins (Sfps) to their mates during copulation
Sfps are thought to interact with several classes of molecules, including other Sfps, proteins native to the female reproductive tract, and pathogens that may be introduced during the course of mating
Some of these proteins show elevated rates of evolution, consistent with their involvement in sexual selection or sexual conflict, and many have arisen by tandem gene duplication
Summary
Males of many internally fertilizing species transfer seminal fluid proteins (Sfps) to their mates during copulation. Sfps are thought to interact with several classes of molecules, including other Sfps (which may originate from seminal fluid of the same male or from a competitor), proteins native to the female reproductive tract, and pathogens that may be introduced during the course of mating. These interactions create opportunities for coevolution, leading to speculation that sperm competition, sexual conflict, sexual selection, and/or host–pathogen interactions could drive the rapid, adaptive evolution of many Sfps [3]. Researchers have sought to identify and characterize Sfps in such diverse taxa as mosquitoes, crickets, honeybees, rodents, and primates [6,7,8,9,10]
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