Context-dependent thermolability of sex determination in a lacertid lizard with heteromorphic sex chromosomes.

Developmental conditions can profoundly impact key life history traits of the individual. In cases where offspring sex is driven by developmental reaction norms, permanent changes to the phenotype can fundamentally alter life history trajectories. Sex determination mechanisms in reptiles are remarkably diverse, including well-characterised genetic and temperature-dependent sex determination. In rarer, but increasingly more commonly documented cases, sex can also be determined by a combination of the two, with temperature overriding the genetically determined sex. Thus, sex-by-temperature interactions is a mechanism that can be contextually labile, where reaction norms of sex against developmental environment might only be observable under certain conditions. We examine the effects of incubation temperature on hatchling sex in an oviparous lizard with clearly defined heteromorphic sex chromosomes presumed to determine sex solely on a genetic basis. We also test the repeatability of our results by replicating incubation experiments across 3 years. We show that warmer temperatures may override chromosomal sex and cause an overproduction of daughters. However, this effect was inconsistent among years, with high temperature only resulting in a daughter-significant bias in one year. Warm-incubated daughters were more efficient at converting yolk into tissue, which would allow for greater resource allocation to other fitness-related processes, such as growth. This suggests that thermolabile sex determination could be a trait under selection. More energy-efficient embryos also produced faster-growing offspring, suggesting that energy utilization patterns of the embryo were maintained into the juvenile stage, which could have important implications for the ontogenetic development and evolution of life histories.

Quantitative genetics of breeding coloration in sand lizards; genic capture unlikely to maintain additive genetic variance.

Sexual selection on fitness-determining traits should theoretically erode genetic variance and lead to low heritability. However, many sexually selected traits maintain significant phenotypic and additive genetic variance, with explanations for this "lek paradox" including genic capture due to condition-dependence, and breaks on directional selection due to environmental sources of variance including maternal effects. Here we investigate genetic and environmental sources of variance in the intrasexually selected green badge of the sand lizard (Lacerta agilis). The badge functions as a cue to male fighting ability in this species, and male-male interactions determine mate acquisition. Using animal models on a pedigree including three generations of males measured over an extensive 9-year field study, we partition phenotypic variance in both badge size and body condition into additive genetic, maternal, and permanent environmental effects experienced by an individual over its lifespan. Heritability of badge size was 0.33 with a significant estimate of underlying additive genetic variance. Body condition was strongly environmentally determined in this species and did not show either significant additive genetic variance or heritability. Neither badge size nor body condition was responsive to maternal effects. We propose that the lack of additive genetic variance and heritability of body condition makes it unlikely that genic capture mechanisms maintain additive genetic variance for badge size. That said, genic capture was originally proposed for male traits under female choice, not agonistic selection. If developmental pathways generating variance in body condition, and/or the covarying secondary sex trait, differ between inter- and intrasexual selection, or the rate at which their additive genetic variance or covariance is depleted, future work may show whether genic capture is largely restricted to intersexual selection processes.

Limited effects of inbreeding on breeding coloration.

Animal color signals may function as indicators of fighting ability when males compete for access to females. This allows opponents to settle aggressive interactions before they escalate into physical combat and injury. Thus, there may be strong directional selection on these traits, toward enhanced signal quality. This renders sexually selected traits particularly susceptible to inbreeding depression, due to relatively low ratios of additive genetic variance to dominance variance. We measured the effects of inbreeding on an intrasexually selected color signal (the badge) in a population of Swedish sand lizards (Lacerta agilis) using the Rhh software based on 17 to 21 microsatellites. Males of this sexually dichromatic species use the badge during aggressive interactions to display, and assess, fighting ability. We found negative effects of homozygosity on badge size, saturation, and brightness. However, no such effects were observed on color hue. Pairwise correlations between badge size, hue, and saturation were all statistically significant. Thus, the sand lizard "badge" is a multicomponent signal with variation explained by covariation in badge size, saturation, and color hue. Body mass corrected for skeletal size (body condition) positively predicted badge size and saturation, encouraging future research on the extent that sexual signals may convey information on multigene targets (i.e. "genic capture").

Seasonal but not sex-biased gene expression of the carotenoid ketolase, CYP2J19, in the sexually dichromatic southern red bishop (Euplectes orix).

Intense red colours in birds are often owing to ketocarotenoids (KCs). In many land birds, KCs are oxidized from dietary yellow precursors, presumably by the avian carotenoid ketolase CYP2J19, the regulation and constraints of which have important implications for condition-dependence and honest signalling of carotenoid colour displays. We investigated hepatic CYP2J19 gene expression in the seasonally and sexually dichromatic southern red bishop (Euplectes orix) in relation to season, sex, progression of the prenuptial moult, testis size, body condition, redness and circulating sex steroids. A coloration function of CYP2J19 is supported by a seasonal upregulation prior to and during the carotenoid-depositing stage of the male prenuptial moult. However, CYP2J19 expression was similarly high in females (which do not moult prenuptially), and remained high in males after moult, suggesting additional or alternative roles of hepatic CYP2J19 or its products, such as detoxification or antioxidant functions. In males, the CYP2J19 upregulation preceded and was unrelated to the rise in plasma testosterone, but was correlated with androstenedione, probably of adrenal origin and compatible with luteinizing hormone-induced and (in females) oestrogen-suppressed moult. Finally, contrary to ideas that carotenoid ketolation rate mediates honest signalling of male quality, CYP2J19 expression was not related to plumage redness or male body condition.

Layered evolution of gene expression in "superfast" muscles for courtship.

Identifying the molecular process of complex trait evolution is a core goal of biology. However, pinpointing the specific context and timing of trait-associated changes within the molecular evolutionary history of an organism remains an elusive goal. We study this topic by exploring the molecular basis of elaborate courtship evolution, which represents an extraordinary example of trait innovation. Within the behaviorally diverse radiation of Central and South American manakin birds, species from two separate lineages beat their wings together using specialized "superfast" muscles to generate a "snap" that helps attract mates. Here, we develop an empirical approach to analyze phylogenetic lineage-specific shifts in gene expression in the key snap-performing muscle and then integrate these findings with comparative transcriptomic sequence analysis. We find that rapid wing displays are associated with changes to a wide range of molecular processes that underlie extreme muscle performance, including changes to calcium trafficking, myocyte homeostasis and metabolism, and hormone action. We furthermore show that these changes occur gradually in a layered manner across the species history, wherein which ancestral genetic changes to many of these molecular systems are built upon by later species-specific shifts that likely finalized the process of display performance adaptation. Our study demonstrates the potential for combining phylogenetic modeling of tissue-specific gene expression shifts with phylogenetic analysis of lineage-specific sequence changes to reveal holistic evolutionary histories of complex traits.

Correlated Evolution of Acrobatic Display and Both Neural and Somatic Phenotypic Traits in Manakins (Pipridae).

Brightly colored manakin (Aves: Pipridae) males are known for performing acrobatic displays punctuated by non-vocal sounds (sonations) in order to attract dull colored females. The complexity of the display sequence and assortment of display elements involved (e.g., sonations, acrobatic maneuvers, and cooperative performances) varies considerably across manakin species. Species-specific display elements coevolve with display-distinct specializations of the neuroanatomical, muscular, endocrine, cardiovascular, and skeletal systems in the handful of species studied. Conducting a broader comparative study, we previously found positive associations between display complexity and both brain mass and body mass across eight manakin genera, indicating selection for neural and somatic expansion to accommodate display elaboration. Whether this gross morphological variation is due to overall brain and body mass expansion (concerted evolution) versus size increases in only functionally relevant brain regions and growth of particular body ("somatic") features (mosaic evolution) remains to be explored. Here, we test the hypothesis that cross-species variation in male brain mass and body mass is driven by mosaic evolution. We predicted positive associations between display complexity and variation in the volume of the cerebellum and sensorimotor arcopallium, brain regions which have roles in sensorimotor processes, and learning and performance of precisely timed and sequenced thoughts and movements, respectively. In contrast, we predicted no associations between the volume of a limbic arcopallial nucleus or a visual thalamic nucleus and display complexity as these regions have no-specific functional relationship to display behavior. For somatic features, we predicted that the relationship between body mass and complexity would not include contributions of tarsus length based on a recent study suggesting selection on tarsus length is less labile than body mass. We tested our hypotheses in males from 12 manakin species and a closely related flycatcher. Our analyses support mosaic evolution of neural and somatic features functionally relevant to display and indicate that sexual selection for acrobatic complexity increases the capacity for procedural learning via cerebellar enlargement and may decrease maneuverability via increases in tarsus length.

Vitellogenin offsets oxidative costs of reproduction in female painted dragon lizards.

Vitellogenesis ('yolking' of follicles) is a bioenergetically costly stage of reproduction requiring enlargement of the liver to produce vitellogenin (VTG) yolk precursor proteins, which are transported and deposited at the ovary. VTG may, however, serve non-nutritive anti-oxidant functions, a hypothesis supported by empirical work on aging and other life-history transitions in several taxa. We test this hypothesis in female painted dragon lizards (Ctenophorus pictus) by examining covariation in VTG with the ovarian cycle, and relative to reactive oxygen species (ROS) including baseline superoxide (bSO). Plasma VTG decreased prior to ovulation, when VTG is deposited into follicles. VTG, however, remained elevated post-ovulation when no longer necessary for yolk provisioning and was unrelated to reproductive investment. Instead, VTG was strongly and positively predicted by prior bSO. ROS, in turn, was negatively predicted by prior VTG, while simultaneously sampled VTG was a positive predictor. These findings are consistent with the hypothesis that VTG functions as an anti-oxidant to counteract oxidative stress associated with vitellogenesis. The relationship between bSO and VTG was strongest in post-ovulatory females, indicating that its function may be largely anti-oxidant at this time. In conclusion, VTG may be under selection to offset oxidative costs of reproduction in egg-producing species.

Endless forms of sexual selection.

In recent years, the field of sexual selection has exploded, with advances in theoretical and empirical research complementing each other in exciting ways. This perspective piece is the product of a "stock-taking" workshop on sexual selection and sexual conflict. Our aim is to identify and deliberate on outstanding questions and to stimulate discussion rather than provide a comprehensive overview of the entire field. These questions are organized into four thematic sections we deem essential to the field. First we focus on the evolution of mate choice and mating systems. Variation in mate quality can generate both competition and choice in the opposite sex, with implications for the evolution of mating systems. Limitations on mate choice may dictate the importance of direct vs. indirect benefits in mating decisions and consequently, mating systems, especially with regard to polyandry. Second, we focus on how sender and receiver mechanisms shape signal design. Mediation of honest signal content likely depends on integration of temporally variable social and physiological costs that are challenging to measure. We view the neuroethology of sensory and cognitive receiver biases as the main key to signal form and the 'aesthetic sense' proposed by Darwin. Since a receiver bias is sufficient to both initiate and drive ornament or armament exaggeration, without a genetically correlated or even coevolving receiver, this may be the appropriate 'null model' of sexual selection. Thirdly, we focus on the genetic architecture of sexually selected traits. Despite advances in modern molecular techniques, the number and identity of genes underlying performance, display and secondary sexual traits remains largely unknown. In-depth investigations into the genetic basis of sexual dimorphism in the context of long-term field studies will reveal constraints and trajectories of sexually selected trait evolution. Finally, we focus on sexual selection and conflict as drivers of speciation. Population divergence and speciation are often influenced by an interplay between sexual and natural selection. The extent to which sexual selection promotes or counteracts population divergence may vary depending on the genetic architecture of traits as well as the covariance between mating competition and local adaptation. Additionally, post-copulatory processes, such as selection against heterospecific sperm, may influence the importance of sexual selection in speciation. We propose that efforts to resolve these four themes can catalyze conceptual progress in the field of sexual selection, and we offer potential avenues of research to advance this progress.

Testosterone activates sexual dimorphism including male-typical carotenoid but not melanin plumage pigmentation in a female bird.

In males it is frequently testosterone (T) that activates the expression of sexually selected morphological and behavioral displays, but the role of T in regulating similar traits in females is less clear. Here, we combine correlational data with results from T and gonadotropin-releasing hormone (GnRH) manipulations in both sexes to assess the role of T in mediating sexually dimorphic coloration and morphology in the red-backed fairy-wren (Malurus melanocephalus). We show that: (1) natural variation in female expression of ornamental traits (darkened bills and red back feathers) is positively associated with age and circulating androgen titres, (2) females have the capacity to express most male-typical traits in response to exogenous T, including carotenoid-pigmented body plumage, shorter feathers, darkened bill and enlarged cloacal protuberance, but (3) appear constrained in production of male-typical melanin-pigmented plumage, and (4) low androgen levels during the pre-nuptial molt, probably because of low ovarian capacity for steroid production (or luteinizing hormone sensitivity), prevent females from developing male-like ornamentation. Thus, females appear to retain molecular mechanisms for hormonally regulated male-typical ornamentation, although these are rarely activated because of insufficient production of the hormonal signal.

Adaptive evolution of a derived radius morphology in manakins (Aves, Pipridae) to support acrobatic display behavior.

The morphology of the avian skeleton is often studied in the context of adaptations for powered flight. The effects of other evolutionary forces, such as sexual selection, on avian skeletal design are unclear, even though birds produce diverse behaviors that undoubtedly require a variety of osteological modifications. Here, we investigate this issue in a family of passerine birds called manakins (Pipridae), which have evolved physically unusual and elaborate courtship displays. We report that, in species within the genus Manacus, the shaft of the radius is heavily flattened and shows substantial solidification. Past work anecdotally notes this morphology and attributes it to the species' ability to hit their wings together above their heads to produce loud mechanical sonations. Our results show that this feature is unique to Manacus compared to the other species in our study, including a variety of taxa that produce other sonations through alternate wing mechanisms. At the same time, our data reveal striking similarities across species in total radius volume and solidification. Together, this suggests that supposedly adaptive alterations in radial morphology occur within a conserved framework of a set radius volume and solidness, which in turn is likely determined by natural selection. Further allometric analyses imply that the radius is less constrained by body size and the structural demands that underlie powered flight, compared to other forelimb bones that are mostly unmodified across taxa. These results are consistent with the idea that the radius is more susceptible to selective modification by sexual selection. Overall, this study provides some of the first insight into the osteological evolution of passerine birds, as well as the way in which opposing selective forces can shape skeletal design in these species. J. Morphol. 277:766-775, 2016. © 2016 Wiley Periodicals, Inc.

Corticosterone: a costly mediator of signal honesty in sand lizards.

The mechanisms underlying honest signal expression remain elusive and may involve the integration of social and physiological costs. Corticosterone is a socially modulated metabolic hormone that mediates energy investment and behavior and may therefore function to deter dishonest signal expression. We examined the relationship between corticosterone and green badge coloration in male sand lizards (Lacerta agilis), hypothesizing that physiological and behavioral costs resulting from elevated baseline glucocorticoids function in maintenance of honest signal expression. We found that large-badged males had higher corticosterone titer, with this relationship apparent at the end of the season and absent early in the season. Large-badged males also suffered higher ectoparasite load (number of tick nymphs), despite being in better condition than small-badged males. Ectoparasite load was positively related to corticosterone titer early in the season at the time of badge formation. High-condition individuals had lower corticosterone and lower numbers of ectoparasites than low-condition individuals, suggestive of conditional variation in ability to withstand costs of corticosterone. We found an opposing negative relationship between corticosterone titer and endoparasite load. Corticosterone titer was also negatively associated with male mobility, a fitness-determining behavior in this species. Because badge size is involved in mediating agonistic social interactions in this species, our results suggest that badge-dependent variation in corticosterone is likely reflective of variation in social conditions experienced over the course of the season. Our results implicate corticosterone in maintenance of signal honesty, both early in the season through enforcement of physiological costs (ectoparasite load) and during the season through behavioral costs (male mobility). We propose that socially modulated variation in corticosterone critically functions in mediation of signal honesty without requiring a direct role for corticosterone in trait expression.

Evolutionary patterns of adaptive acrobatics and physical performance predict expression profiles of androgen receptor - but not oestrogen receptor - in the forelimb musculature.

1. Superior physical competence is vital to the adaptive behavioral routines of many animals, particularly those that engage in elaborate socio-sexual displays. How such traits evolve across species remains unclear. 2. Recent work suggests that activation of sex steroid receptors in neuromuscular systems is necessary for the fine motor skills needed to execute physically elaborate displays. Thus, using passerine birds as models, we test whether interspecific variation in display complexity predicts species differences in the abundance of androgen and estrogen receptors (AR and ERα) expressed in the forelimb musculature and spinal cord. 3. We find that small-scale evolutionary patterns in physical display complexity positively predict expression of the AR in the main muscles that lift and retract the wings. No such relationship is detected in the spinal cord, and we do not find a correlation between display behavior and neuromuscular expression of ERα. Also, we find that AR expression levels in different androgen targets throughout the body - namely the wing muscles, spinal cord, and testes - are not necessarily correlated, providing evidence that evolutionary forces may drive AR expression in a tissue-specific manner. 4. These results suggest co-evolution between the physical prowess necessary for display performance and levels of AR expression in avian forelimb muscles. Moreover, this relationship appears to be specific to muscle and AR-mediated, but not ERα-mediated, signaling. 5. Given that prior work suggests that activation of muscular AR is a necessary component of physical display performance, our current data support the hypothesis that sexual selection shapes levels of AR expressed in the forelimb skeletal muscles to help drive the evolution of adaptive motor abilities.

Acrobatic courtship display coevolves with brain size in manakins (Pipridae).

Acrobatic display behaviour is sexually selected in manakins (Pipridae) and can place high demands on many neural systems. Manakin displays vary across species in terms of behavioural complexity, differing in number of unique motor elements, production of mechanical sounds, cooperation between displaying males, and construction of the display site. Historically, research emphasis has been placed on neurological specializations for vocal aspects of courtship, and less is known about the control of physical, non-vocal displays. By examining brain evolution in relation to extreme acrobatic feats such as manakin displays, we can vastly expand our knowledge of how sexual selection acts on motor behaviour. We tested the hypothesis that sexual selection for complex motor displays has selected for larger brains across the Pipridae. We found that display complexity positively predicts relative brain weight (adjusted for body size) after controlling for phylogeny in 12 manakin species and a closely related flycatcher. This evidence suggests that brain size has evolved in response to sexual selection to facilitate aspects of display such as motor, sensorimotor, perceptual, and cognitive abilities. We show, for the first time, that sexual selection for acrobatic motor behaviour can drive brain size evolution in avian species and, in particular, a family of suboscine birds.

Sexually selected male plumage color is testosterone dependent in a tropical passerine bird, the red-backed fairy-wren (Malurus melanocephalus).

BACKGROUND: Sexual signals, such as bright plumage coloration in passerine birds, reflect individual quality, and testosterone (T) may play a critical role in maintaining signal honesty. Manipulations of T during molt have yielded mixed effects on passerine plumage color, in most cases delaying molt or leading to production of drab plumage. However, the majority of these studies have been conducted on species that undergo a post-nuptial molt when T is low; the role of T in species that acquire breeding plumage during a pre-nuptial molt remains largely unexplored. METHODOLOGY/PRINCIPAL FINDINGS: We experimentally tested the effects of increased T on plumage color in second-year male red-backed fairy-wrens (Malurus melanocephalus), a species in which after-second-year males undergo a pre-nuptial molt into red/black (carotenoid and melanin-based) plumage and second-year males either assume red/black or brown breeding plumage. T treatment stimulated a rapid and early onset pre-nuptial molt and resulted in red/black plumage acquisition, bill darkening, and growth of the sperm storage organ, but had no effect on body condition or corticosterone concentrations. Control males molted later and assumed brown plumage. T treated males produced feathers with similar but not identical reflectance parameters to those of unmanipulated after-second-year red/black males; while reflectance spectra of red back and black crown feathers were similar, black breast feathers differed in UV chroma, hue and brightness, indicating a potentially age and plumage patch-dependent response to T for melanin- vs. carotenoid-pigmentation. CONCLUSIONS/SIGNIFICANCE: We show that testosterone is the primary mechanism functioning during the pre-nuptial molt to regulate intrasexually variable plumage color and breeding phenotype in male red-backed fairy-wrens. Our results suggest that the effects of T on plumage coloration may vary with timing of molt (pre- vs. post-nuptial), and that the role of T in mediating plumage signal production may differ across age classes, plumage patches, and between pigment-types.