Abstract

The seminal fluid proteins (SFPs) transferred to mating partners along with sperm often play crucial roles in mediating post-mating sexual selection. One way in which sperm donors can maximize their own reproductive success is by modifying the partner's (sperm recipient's) post-copulatory behaviour to prevent or delay re-mating, thereby decreasing the likelihood or intensity of sperm competition. Here, we adopted a quantitative genetic approach combining gene expression and behavioural data to identify candidates that could mediate such a response in the simultaneously hermaphroditic flatworm Macrostomum lignano. We identified two putative SFPs-Mlig-pro46 and Mlig-pro63-linked to both mating frequency and 'suck' frequency, a distinctive behaviour, in which, upon ejaculate receipt, the worm places its pharynx over its female genital opening and apparently attempts to remove the received ejaculate. We, therefore, performed a manipulative experiment using RNA interference-induced knockdown to ask how the loss of Mlig-pro46 and Mlig-pro63 expression, singly and in combination, affects mating frequency, partner suck propensity and sperm competitive ability. None of the knockdown treatments impacted strongly on the mating frequency or sperm competitive ability, but knockdown of Mlig-pro63 resulted in a significantly decreased suck propensity of mating partners. This suggests that Mlig-pro63 may normally act as a cue in the ejaculate to trigger recipient suck behaviour and-given that other proteins in the ejaculate have the opposite effect-could be one component of an ongoing arms race between donors and recipients over the control of ejaculate fate. However, the adaptive significance of Mlig-pro46 and Mlig-pro63 from a donor perspective remains enigmatic.

Highlights

  • In polyandrous species, if females store sperm from multiple males, post-mating sexual selection can occur when sperm from different males compete with each other for fertilization and/or when females choose specific sperm to fertilize eggs

  • The seminal fluid proteins (SFPs) found in the ejaculate play crucial roles in reproduction and can modulate the mating partner's behaviour and physiology such that they affect the reproductive success of both partners, making these proteins key mediators of post-mating sexual selection (Cameron, Day, & Rowe, 2007; Chapman, 2001; Hodgson & Hosken, 2006; Poiani, 2006; Ram & Wolfner, 2007a)

  • Because the same two candidate transcripts, Mlig-pro46 and Mlig-pro63, were positively loaded on an additional axis of seminal fluid variation, PC3, and this was positively correlated with suck propensity in the pairs environment (Patlar et al, 2020), we further investigated whether RNAi knockdown impacted on partner suck behaviour

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Summary

| INTRODUCTION

If females store sperm from multiple males, post-mating sexual selection can occur when sperm from different males compete with each other for fertilization (i.e. sperm competition; Parker, 1970) and/or when females choose specific sperm to fertilize eggs (i.e. cryptic female choice; Eberhard,1996; Thornhill, 1983). The intravaginal injection of one L. stagnalis SFP (LyAcp10) led to a decrease in egg laying (Koene et al, 2010), and the injection of two other SFPs (LyAcp8b and LyAcp5) reduced the number of sperm transferred by the recipient in a subsequent mating as a donor and, as a result, decreased their paternity success (Nakadera et al, 2014; see Schärer, 2014) This latter effect emphasizes that it is a potentially adaptive strategy in simultaneous hermaphrodites to steer your partner away from its male function and that the action of seminal fluid may be one means of doing so (Schärer & Ramm, 2016). Both putative SFPs were already included in the above-mentioned naïve RNAi screen (Weber et al, 2019), in which Mlig-pro showed some evidence for a reduced offensive sperm competitive ability (P2, see below), making it an even more promising candidate for more detailed characterization

| MATERIALS AND METHODS
Findings
| DISCUSSION
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