Dynamic digestive physiology of a female reproductive organ in a polyandrous butterfly

Abstract

Reproductive traits experience high levels of selection because of their direct ties to fitness, often resulting in rapid adaptive evolution. Much of the work in this area has focused on male reproductive traits. However, a more comprehensive understanding of female reproductive adaptations and their relationship to male characters is crucial to uncover the relative roles of sexual cooperation and conflict in driving co-evolutionary dynamics between the sexes. We focus on the physiology of a complex female reproductive adaptation in butterflies and moths: a stomach-like organ in the female reproductive tract called the bursa copulatrix that digests the male ejaculate (spermatophore). Little is known about how the bursa digests the spermatophore. We characterized bursa proteolytic capacity in relation to female state in the polyandrous butterfly Pieris rapae. We found that the virgin bursa exhibits extremely high levels of proteolytic activity. Furthermore, in virgin females, bursal proteolytic capacity increases with time since eclosion and ambient temperature, but is not sensitive to the pre-mating social environment. Post copulation, bursal proteolytic activity decreases rapidly before rebounding toward the end of a mating cycle, suggesting active female regulation of proteolysis and/or potential quenching of proteolysis by male ejaculate constituents. Using transcriptomic and proteomic approaches, we report identities for nine proteases actively transcribed by bursal tissue and/or expressed in the bursal lumen that may contribute to observed bursal proteolysis. We discuss how these dynamic physiological characteristics may function as female adaptations resulting from sexual conflict over female remating rate in this polyandrous butterfly.

Structural complexity and molecular heterogeneity of a butterfly ejaculate reflect a complex history of selection

“…Because the spermatophores sampled here were collected 2 h after mating-the earliest time point at which the envelope is fully hardened-these proteases could have been incorporated either during the transfer of the ejaculate or shortly after the transfer was complete. This rapid incorporation of large amounts of female-derived proteases is consistent with our prior observations of the remarkably high concentrations of bursal proteases in virgin females (34,38) and implies that females may already have an active role in spermatophore digestion even before breach of the indigestible envelope by the signum.…”

“…We found large amounts of female-produced proteases throughout the spermatophore, making up 25% and 11% of soluble protein in the envelope and inner matrix, respectively. These proteases are expressed at extraordinary levels in virgin females (38), suggesting that females may concentrate them in the bursa and/or reproductive tract before mating in an attempt to incorporate them into the spermatophore either during formation or shortly thereafter. These proteases may then increase the rate of digestion of the spermatophore material, particularly the inner matrix, and may also assist the signum in breaching the outer envelope.…”

“…1A). Interestingly, all six of these female-produced proteins are proteases, each of which was previously identified as expressed in the bursa copulatrix (38). Because the spermatophores sampled here were collected 2 h after mating-the earliest time point at which the envelope is fully hardened-these proteases could have been incorporated either during the transfer of the ejaculate or shortly after the transfer was complete.…”

“…Muscular contractions of the bursal wall animate a hardened, toothed device called the signum, which bores a hole through the tough outer envelope (34,36,37). Chemical breakdown of the spermatophore is then accomplished through a mixture of enzymes, including as many as nine proteases, which the female secretes in concentrations dramatically surpassing those found in the larval gut (38).…”

Seminal fluid protein divergence among populations exhibiting postmating prezygotic reproductive isolation

“…Proteases and peptidases are central to reproduction across taxa, regulating proteolytic activity and initiating cascades of interactions among downstream proteins (Bayram et al, 2017(Bayram et al, , 2019LaFlamme et al, 2012;LaFlamme & Wolfner, 2013;Plakke et al, 2015Plakke et al, , 2019.…”