Assessing the evolutionary lability of insulin signalling in the regulation of nutritional plasticity across traits and species of horned dung beetles.

Nutrition-dependent growth of sexual traits is a major contributor to phenotypic diversity, and a large body of research documents insulin signalling as a major regulator of nutritional plasticity. However, findings across studies raise the possibility that the role of individual components within the insulin signalling pathway diverges in function among traits and taxa. Here, we use RNAi-mediated transcript depletion in the gazelle dung beetle to investigate the functions of forkhead box O (Foxo) and two paralogs of the insulin receptor (InR1 and InR2) in shaping nutritional plasticity in polyphenic male head horns, exaggerated fore legs, and weakly nutrition-responsive genitalia. Our functional genetic manipulations led to three main findings: FoxoRNAi reduced the length of exaggerated head horns in large males, while neither InR1 nor InR2 knock-downs resulted in measurable horn phenotypes. These results are similar to those documented previously for another dung beetle (Onthophagus taurus), but in stark contrast to findings in rhinoceros beetles. Secondly, knockdown of Foxo, InR1, and InR2 led to an increase in the intercept or slope of the scaling relationship of genitalia size. These findings are in contrast even to results documented previously for O. taurus. Lastly, while FoxoRNAi reduces male forelegs in D. gazella and O. taurus, the effects of InR1 and InR2 knockdowns diverged across dung beetle species. Our results add to the growing body of literature indicating that despite insulin signalling's conserved role as a regulator of nutritional plasticity, the functions of its components may diversify among traits and species, potentially fuelling the evolution of scaling relationships.

A Mediator subunit imparts robustness to a polyphenism decision.

Polyphenism is a type of developmental plasticity that translates continuous environmental variability into discontinuous phenotypes. Such discontinuity likely requires a switch between alternative gene-regulatory networks, a principle that has been borne out by mechanisms found to promote morph-specific gene expression. However, whether robustness is required to execute a polyphenism decision has awaited testing at the molecular level. Here, we used a nematode model for polyphenism, Pristionchus pacificus, to identify the molecular regulatory factors that ensure the development of alternative forms. This species has a dimorphism in its adult feeding structures, specifically teeth, which are a morphological novelty that allows predation on other nematodes. Through a forward genetic screen, we determined that a duplicate homolog of the Mediator subunit MDT-15/MED15, P. pacificus MDT-15.1, is necessary for the polyphenism and the robustness of the resulting phenotypes. This transcriptional coregulator, which has a conserved role in metabolic responses to nutritional stress, coordinates these processes with its effects on this diet-induced polyphenism. Moreover, this MED15 homolog genetically interacts with two nuclear receptors, NHR-1 and NHR-40, to achieve dimorphism: Single and double mutants for these three factors result in morphologies that together produce a continuum of forms between the extremes of the polyphenism. In summary, we have identified a molecular regulator that confers discontinuity to a morphological polyphenism, while also identifying a role for MED15 as a plasticity effector.

Polyphenism of a Novel Trait Integrated Rapidly Evolving Genes into Ancestrally Plastic Networks.

Developmental polyphenism, the ability to switch between phenotypes in response to environmental variation, involves the alternating activation of environmentally sensitive genes. Consequently, to understand how a polyphenic response evolves requires a comparative analysis of the components that make up environmentally sensitive networks. Here, we inferred coexpression networks for a morphological polyphenism, the feeding-structure dimorphism of the nematode Pristionchus pacificus. In this species, individuals produce alternative forms of a novel trait-moveable teeth, which in one morph enable predatory feeding-in response to environmental cues. To identify the origins of polyphenism network components, we independently inferred coexpression modules for more conserved transcriptional responses, including in an ancestrally nonpolyphenic nematode species. Further, through genome-wide analyses of these components across the nematode family (Diplogastridae) in which the polyphenism arose, we reconstructed how network components have changed. To achieve this, we assembled and resolved the phylogenetic context for five genomes of species representing the breadth of Diplogastridae and a hypothesized outgroup. We found that gene networks instructing alternative forms arose from ancestral plastic responses to environment, specifically starvation-induced metabolism and the formation of a conserved diapause (dauer) stage. Moreover, loci from rapidly evolving gene families were integrated into these networks with higher connectivity than throughout the rest of the P. pacificus transcriptome. In summary, we show that the modular regulatory outputs of a polyphenic response evolved through the integration of conserved plastic responses into networks with genes of high evolutionary turnover.

Nutrition-responsive gene expression and the developmental evolution of insect polyphenism.

Nutrition-responsive development is a ubiquitous and highly diversified example of phenotypic plasticity, yet its underlying molecular and developmental mechanisms and modes of evolutionary diversification remain poorly understood. We measured genome-wide transcription in three closely related species of horned beetles exhibiting strikingly diverse degrees of nutrition responsiveness in the development of male weaponry. We show that (1) counts of differentially expressed genes between low- and high-nutritional backgrounds mirror species-specific degrees of morphological nutrition responsiveness; (2) evolutionary exaggeration of morphological responsiveness is underlain by both amplification of ancestral nutrition-responsive gene expression and recruitment of formerly low nutritionally responsive genes; and (3) secondary loss of morphological responsiveness to nutrition coincides with a dramatic reduction in gene expression plasticity. Our results further implicate genetic accommodation of ancestrally high variability of gene expression plasticity in both exaggeration and loss of nutritional plasticity, yet reject a major role of taxon-restricted genes in the developmental regulation and evolution of nutritional plasticity.

Developmental bias in horned dung beetles and its contributions to innovation, adaptation, and resilience.

Developmental processes transduce diverse influences during phenotype formation, thereby biasing and structuring amount and type of phenotypic variation available for evolutionary processes to act on. The causes, extent, and consequences of this bias are subject to significant debate. Here we explore the role of developmental bias in contributing to organisms' ability to innovate, to adapt to novel or stressful conditions, and to generate well integrated, resilient phenotypes in the face of perturbations. We focus our inquiry on one taxon, the horned dung beetle genus Onthophagus, and review the role developmental bias might play across several levels of biological organization: (a) gene regulatory networks that pattern specific body regions; (b) plastic developmental mechanisms that coordinate body wide responses to changing environments and; (c) developmental symbioses and niche construction that enable organisms to build teams and to actively modify their own selective environments. We posit that across all these levels developmental bias shapes the way living systems innovate, adapt, and withstand stress, in ways that can alternately limit, bias, or facilitate developmental evolution. We conclude that the structuring contribution of developmental bias in evolution deserves further study to better understand why and how developmental evolution unfolds the way it does.

Evolution of, and via, Developmental Plasticity: Insights through the Study of Scaling Relationships.

Scaling relationships emerge from differential growth of body parts relative to each other. As such, scaling relationships are at least in part the product of developmental plasticity. While some of the developmental genetic mechanisms underlying scaling relationships are starting to be elucidated, how these mechanisms evolve and give rise to the enormous diversity of allometric scaling observed in nature is less understood. Furthermore, developmental plasticity has itself been proposed as a mechanism that facilitates adaptation and diversification, yet its role in the developmental evolution of scaling relationships remains largely unknown. In this review, we first explore how the mechanisms of scaling relationships have evolved. We primarily focus on insect development and review how pathway components and pathway interactions have evolved across taxa to regulate scaling relationships across diverse traits. We then discuss the potential role of developmental plasticity in the evolution of scaling relationships. Specifically, we address the potential role of allometric plasticity and cryptic genetic variation in allometry in facilitating divergence via genetic accommodation. Collectively, in this article, we aim to bring together two aspects of developmental plasticity: the mechanistic underpinnings of scaling relationships and their evolution, and the potential role that plasticity plays in the evolutionary diversification of scaling relationships.

The sequencing and interpretation of the genome obtained from a Serbian individual.

Recent genetic studies and whole-genome sequencing projects have greatly improved our understanding of human variation and clinically actionable genetic information. Smaller ethnic populations, however, remain underrepresented in both individual and large-scale sequencing efforts and hence present an opportunity to discover new variants of biomedical and demographic significance. This report describes the sequencing and analysis of a genome obtained from an individual of Serbian origin, introducing tens of thousands of previously unknown variants to the currently available pool. Ancestry analysis places this individual in close proximity to Central and Eastern European populations; i.e., closest to Croatian, Bulgarian and Hungarian individuals and, in terms of other Europeans, furthest from Ashkenazi Jewish, Spanish, Sicilian and Baltic individuals. Our analysis confirmed gene flow between Neanderthal and ancestral pan-European populations, with similar contributions to the Serbian genome as those observed in other European groups. Finally, to assess the burden of potentially disease-causing/clinically relevant variation in the sequenced genome, we utilized manually curated genotype-phenotype association databases and variant-effect predictors. We identified several variants that have previously been associated with severe early-onset disease that is not evident in the proband, as well as putatively impactful variants that could yet prove to be clinically relevant to the proband over the next decades. The presence of numerous private and low-frequency variants, along with the observed and predicted disease-causing mutations in this genome, exemplify some of the global challenges of genome interpretation, especially in the context of under-studied ethnic groups.

On the Reciprocally Causal and Constructive Nature of Developmental Plasticity and Robustness.

Exposure to environmental variation is a characteristic feature of normal development, one that organisms can respond to during their lifetimes by actively adjusting or maintaining their phenotype in order to maximize fitness. Plasticity and robustness have historically been studied by evolutionary biologists through quantitative genetic and reaction norm approaches, while more recent efforts emerging from evolutionary developmental biology have begun to characterize the molecular and developmental genetic underpinnings of both plastic and robust trait formation. In this review, we explore how our growing mechanistic understanding of plasticity and robustness is beginning to force a revision of our perception of both phenomena, away from our conventional view of plasticity and robustness as opposites along a continuum and toward a framework that emphasizes their reciprocal, constructive, and integrative nature. We do so in three sections. Following an introduction, the first section looks inward and reviews the genetic, epigenetic, and developmental mechanisms that enable organisms to sense and respond to environmental conditions, maintaining and adjusting trait formation in the process. In the second section, we change perspective and look outward, exploring the ways in which organisms reciprocally shape their environments in ways that influence trait formation, and do so through the lens of behavioral plasticity, niche construction, and host-microbiota interactions. In the final section, we revisit established plasticity and robustness concepts in light of these findings, and highlight research opportunities to further advance our understanding of the causes, mechanisms, and consequences of these ubiquitous, and interrelated, phenomena.

Insulin signalling's role in mediating tissue-specific nutritional plasticity and robustness in the horn-polyphenic beetle Onthophagus taurus.

Organisms cope with nutritional variation via developmental plasticity, adjusting trait size to nutrient availability for some traits while enabling others to develop in a nutritionally robust manner. Yet, the developmental mechanisms that regulate organ-specific growth across nutritional gradients remain poorly understood. We assessed the functions of members of the insulin/insulin-like signalling pathway (IIS) in the regulation of nutrition sensitivity and robustness in males of the horn-polyphenic beetle Onthophagus taurus, as well as potential regulatory interactions between IIS and two other growth-regulating pathways: Doublesex and Hedgehog signalling. Using RNA interference (RNAi), we experimentally knocked down both insulin receptors ( InR1 and InR2) and Foxo, a growth inhibitor. We then performed morphometric measurements on horns, a highly nutrition-sensitive trait, and genitalia, a largely nutrition-insensitive trait. Finally, we used quantitative real-time polymerase chain reaction to assess expression levels of doublesex and the Hedgehog signalling gene smoothened following IIS-RNAi. Our results suggest that nutrition responsiveness of both traits is regulated by different IIS components, which transduce nutritional conditions to both Doublesex and Hedgehog pathways, albeit via different IIS pathway members. Combined with previous studies, our findings suggest that separate origins of trait exaggeration among insect lineages were enabled through the independent co-option of IIS, yet via reliance on different components therein.

Evidence of developmental niche construction in dung beetles: effects on growth, scaling and reproductive success.

Niche construction occurs when organisms modify their environments and alter selective conditions through their physiology and behaviours. Such modifications can bias phenotypic variation and enhance organism-environment fit. Yet few studies exist that experimentally assess the degree to which environmental modifications shape developmental and fitness outcomes, how their influences may differ among species and identify the underlying proximate mechanisms. Here, we experimentally eliminate environmental modifications from the developmental environment of Onthophagus dung beetles. We show that these modifications (1) differentially influence growth among species, (2) consistently shape scaling relationships in fitness-related traits, (3) are necessary for the maintenance of sexual dimorphism, (4) influence reproductive success among females of at least one species and (5) implicate larval cultivation of an external rumen as a possible mechanism for environmental modification. Our results present evidence that Onthophagus larvae engage in niche construction, and that this is a fundamental component of beetle development and fitness.

Developmental regulation and evolution of scaling: novel insights through the study of Onthophagus beetles.

Scaling relationships play critical roles in defining biological shape, trait functionality, and species characteristics, yet the developmental basis of scaling and its evolution remain poorly resolved in most taxa. In the horned beetle genus Onthophagus, scaling relationships of most traits are largely comparable across many species, however, the morphology and scaling of horns, a recent evolutionary invention, has diversified dramatically, ranging from modestly to highly positively linear to more complex sigmoidal allometries. Through a series of transcriptomic screens and gene function assays, the doublesex, hedgehog, insulin, and serotonin signaling pathways have recently been implicated in the regulation of amplitude, slope, and threshold location of the highly sigmoidal horn allometry in O. taurus. These and other findings suggest that co-option of these pathways into the regulation of horn development may have been critical in the evolutionary transitions from isometric to positively allometric to sigmoidal allometries in Onthophagus, thereby contributing to the extraordinary diversification of one of the most species-rich genera in the animal kingdom.

Evolutionary Developmental Biology (Evo-Devo) Research in Latin America.

Famous for its blind cavefish and Darwin's finches, Latin America is home to some of the richest biodiversity hotspots of our planet. The Latin American fauna and flora inspired and captivated naturalists from the nineteenth and twentieth centuries, including such notable pioneers such as Fritz Müller, Florentino Ameghino, and Léon Croizat who made a significant contribution to the study of embryology and evolutionary thinking. But, what are the historical and present contributions of the Latin American scientific community to Evo-Devo? Here, we provide the first comprehensive overview of the Evo-Devo laboratories based in Latin America and describe current lines of research based on endemic species, focusing on body plans and patterning, systematics, physiology, computational modeling approaches, ecology, and domestication. Literature searches reveal that Evo-Devo in Latin America is still in its early days; while showing encouraging indicators of productivity, it has not stabilized yet, because it relies on few and sparsely distributed laboratories. Coping with the rapid changes in national scientific policies and contributing to solve social and health issues specific to each region are among the main challenges faced by Latin American researchers. The 2015 inaugural meeting of the Pan-American Society for Evolutionary Developmental Biology played a pivotal role in bringing together Latin American researchers eager to initiate and consolidate regional and worldwide collaborative networks. Such networks will undoubtedly advance research on the extremely high genetic and phenotypic biodiversity of Latin America, bound to be an almost infinite source of amazement and fascinating findings for the Evo-Devo community.