A derived Bmp4 domain causes apoptosis and regression of the embryonic chick phallus

During bird evolution, the phallus was reduced or lost in most lineages

Our results indicate that evolutionary reduction of the intromittent phallus in galliform birds occurred not by disruption of outgrowth signals but by de novo activation of cell death by Bmp4 in the genital tubercle. These findings, together with discoveries implicating Bmps in evolution of beak shape, feathers, and toothlessness, suggest that modulation of Bmp gene regulation played a major role in the evolution of avian morphology.

We investigated genital development in two sister clades of birds, Galliformes (land fowl), most of which lack an intromittent phallus, and Anseriformes (waterfowl), which have well developed phalluses; and in two outgroups, Paleognathae (emus) and Crocodilia (alligators). Galliform embryos undergo cryptic development of a genital tubercle, the precursor of the phallus, but this later undergoes apoptosis, leading to regression of the tubercle. At the molecular level, a derived pattern of Bmp4 expression was identified in chick (a galliform) genital tubercles. Inhibition of Bmp signaling in chick genitalia rescues cells from apoptosis and prevents phallus regression, whereas activation of Bmp signaling in duck (an anseriform) genitalia induces a galliform-like pattern of apoptosis. Thus, distal Bmp activity is necessary and sufficient to induce apoptosis in Galloanserae genital tubercles.

One of the most puzzling events in evolution is the reduction and loss of the phallus in birds. All birds reproduce by internal fertilization, but only ∼3% of birds have retained a phallus capable of intromission. A number of hypotheses have been proposed for the evolutionary mechanisms that drove phallus reduction; however, the underlying developmental mechanisms are unknown.

Introduction

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Briskie J.V. Anatomy and evolution of copulatory structures. Some of the most striking examples of the evolutionary diversification of genital form and function are found in birds []. All birds reproduce by internal fertilization, yet only ∼3% of extant birds possess a phallus capable of intromission []. Indeed, external genitalia are reduced or absent in nearly 10,000 species of birds. These evolutionary changes have resulted in phallus morphologies that can be classified operationally into three general categories: (1) an intromittent phallus that is capable of insertion during copulation (e.g., ducks), (2) a nonintromittent phallus that is reduced but not lost (e.g., chickens), and (3) absence of the phallus that involves a complete loss of a phallic protuberance (e.g., neoaves). Recent progress in phylogenomics has begun to clarify the phylogenetic relationships among birds [], and evolutionary reductions of the intromittent phallus can be mapped to a relatively small number of nodes in the avian phylogeny; for example, after their divergence from Anseriformes (e.g., ducks, swans, geese, and allies), Galliformes (e.g., chickens, quails, pheasants, and megapodes) evolved a highly reduced, nonintromittent phallic rudiment. Even more extreme reduction occurred near the base of Neoaves, the largest clade of birds, which have no remnant of the phallus [].

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Prum R.O. The erection mechanism of the ratite penis. Although most galliforms underwent extreme reduction of the external genitalia [], members of their sister clade, the anseriforms, retained an intromittent phallus, and in some species these elongated coiled organs can exceed the length of the body []. These morphological changes led to changes in copulation strategies in different lineages. For example, male birds that lack a prominent intromittent organ (IO) transfer sperm by apposing the cloaca with that of the female, in a maneuver known as the “cloacal kiss,” which requires cooperation of the female []. By contrast, males with an IO can manipulate females and even forcibly copulate with unwilling females, a behavior that is well documented in waterfowl [].

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Stockley P. Sexual selection and genital evolution. 6 Eberhardt W.G. Sexual Selection and Animal Genitalia. 7 Masly J.P. 170 years of “lock-and-key”: genital morphology and reproductive isolation. 9 Mayr E. Animal Species and Evolution. 2 Hosken D.J.

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Danielsson I. Copulatory behavior, genital morphology, and male fertilization success in water striders. While selective pressures for elaborate phallus morphologies are obvious, it is less clear how selection could favor reduction or loss of an IO. A number of hypotheses have been proposed for the adaptive significance of genital reduction, although the precise nature of the evolutionary mechanisms is unclear due to the paucity of ecological or experimental data []. One of the most influential hypotheses for genital diversity, the “lock and key” hypothesis, argues that rapid evolution of genitalia facilitated speciation through breeding incompatibility (i.e., only males and females of the same species can fit their genitalia together) []. Although anatomical barriers to hybridization can in theory lead to reproductive isolation, experimental studies have failed to support the idea that strict mechanical incompatibility has been a driving force in the evolution of genital form []. Alternatively, Mayr proposed that genital morphology undergoes frequent changes due to pleiotropic effects []. It is unclear why neutral changes resulting from pleiotropy would affect genitalia disproportionately [], although studies of the genetic control of genital development have identified a multitude of developmental mechanisms (including cis-acting DNA regulatory elements) that are shared by other organ systems [], which raises the possibility that modulating gene expression in structures such as limbs could have collateral effects on the genitalia [] A third hypothesis argues that diversification of external genitalia has been driven by sexual selection acting on variation that affects sperm competition, sensory features of genital morphology, female choice, and sexually antagonistic coevolution (i.e., males and females evolve adaptations and counteradaptations to gain control over copulation and fertilization) [].