Experimental evolution of hybrid populations to identify Dobzhansky-Muller incompatibility loci.

Epistatic interactions between loci that reduce fitness in interspecies hybrids, Dobzhansky-Muller incompatibilities (DMIs), contribute genetically to the inviability and infertility within hybrid populations. It remains a challenge, however, to identify the loci that contribute to DMIs as causes of reproductive isolation between species. Here, we assess through forward simulation the power of evolve-and-resequence (E&R) experimental evolution of hybrid populations to map DMI loci. We document conditions under which such a mapping strategy may be most feasible and demonstrate how mapping power is sensitive to biologically relevant parameters such as one-way versus two-way incompatibility type, selection strength, recombination rate, and dominance interactions. We also assess the influence of parameters under direct control of an experimenter, including duration of experimental evolution and number of replicate populations. We conclude that an E&R strategy for mapping DMI loci, and other cases of epistasis, can be a viable option under some circumstances for study systems with short generation times like Caenorhabditis nematodes.

Sexual conflict, heterochrony and tissue specificity as evolutionary problems of adaptive plasticity in development.

Differential gene expression represents a fundamental cause and manifestation of phenotypic plasticity. Adaptive phenotypic plasticity in gene expression as a trait evolves when alleles that mediate gene regulation serve to increase organismal fitness by improving the alignment of variation in gene expression with variation in circumstances. Among the diverse circumstances that a gene encounters are distinct cell types, developmental stages and sexes, as well as an organism's extrinsic ecological environments. Consequently, adaptive phenotypic plasticity provides a common framework to consider diverse evolutionary problems by considering the shared implications of alleles that produce context-dependent gene expression. From this perspective, adaptive plasticity represents an evolutionary resolution to conflicts of interest that arise from any negatively pleiotropic effects of expression of a gene across ontogeny, among tissues, between the sexes, or across extrinsic environments. This view highlights shared properties within the general relation of fitness, trait expression and context that may nonetheless differ substantively in the grain of selection within and among generations to influence the likelihood of adaptive plasticity as an evolutionary response. Research programmes that historically have focused on these separate issues may use the insights from one another by recognizing their shared dependence on context-dependent gene regulatory evolution.

Genomic diversity landscapes in outcrossing and selfing Caenorhabditis nematodes.

Caenorhabditis nematodes form an excellent model for studying how the mode of reproduction affects genetic diversity, as some species reproduce via outcrossing whereas others can self-fertilize. Currently, chromosome-level patterns of diversity and recombination are only available for self-reproducing Caenorhabditis, making the generality of genomic patterns across the genus unclear given the profound potential influence of reproductive mode. Here we present a whole-genome diversity landscape, coupled with a new genetic map, for the outcrossing nematode C. remanei. We demonstrate that the genomic distribution of recombination in C. remanei, like the model nematode C. elegans, shows high recombination rates on chromosome arms and low rates toward the central regions. Patterns of genetic variation across the genome are also similar between these species, but differ dramatically in scale, being tenfold greater for C. remanei. Historical reconstructions of variation in effective population size over the past million generations echo this difference in polymorphism. Evolutionary simulations demonstrate how selection, recombination, mutation, and selfing shape variation along the genome, and that multiple drivers can produce patterns similar to those observed in natural populations. The results illustrate how genome organization and selection play a crucial role in shaping the genomic pattern of diversity whereas demographic processes scale the level of diversity across the genome as a whole.

Speciation and development.

Understanding general principles about the origin of species remains one of the foundational challenges in evolutionary biology. The genomic divergence between groups of individuals can spawn hybrid inviability and hybrid sterility, which presents a tantalizing developmental problem. Divergent developmental programs may yield either conserved or divergent phenotypes relative to ancestral traits, both of which can be responsible for reproductive isolation during the speciation process. The genetic mechanisms of developmental evolution involve cis- and trans-acting gene regulatory change, protein-protein interactions, genetic network structures, dosage, and epigenetic regulation, all of which also have roots in population genetic and molecular evolutionary processes. Toward the goal of demystifying Darwin's "mystery of mysteries," this review integrates microevolutionary concepts of genetic change with principles of organismal development, establishing explicit links between population genetic process and developmental mechanisms in the production of macroevolutionary pattern. This integration aims to establish a more unified view of speciation that binds process and mechanism.

Guerrilla eugenics: gene drives in heritable human genome editing.

CRISPR-Cas9 genome editing can and has altered human genomes, bringing bioethical debates about this capability to the forefront of philosophical and policy considerations. Here, I consider the underexplored implications of CRISPR-Cas9 gene drives for heritable human genome editing. Modification gene drives applied to heritable human genome editing would introduce a novel form of involuntary eugenic practice that I term guerrilla eugenics. Once introduced into a genome, stealth genetic editing by a gene drive genetic element would occur each subsequent generation irrespective of whether reproductive partners consent to it and irrespective of whether the genetic change confers any benefit. By overriding the ability to 'opt in' to genome editing, gene drives compromise the autonomy of carrier individuals and their reproductive partners to choose to use or avoid genome editing and impose additional burdens on those who hope to 'opt out' of further genome editing. High incidence of an initially rare gene drive in small human communities could occur within 200 years, with evolutionary fixation globally in a timeframe that is thousands of times sooner than achievable by non-drive germline editing. Following any introduction of heritable gene drives into human genomes, practices intended for surveillance or reversal also create fundamental ethical problems. Current policy guidelines do not comment explicitly on gene drives in humans. These considerations motivate an explicit moratorium as being warranted on gene drive development in heritable human genome editing.

Regulatory Divergence as a Mechanism for X-Autosome Incompatibilities in Caenorhabditis Nematodes.

The world's astounding biodiversity results from speciation, the process of formation of distinct species. Hybrids between species often have reduced fitness due to negative epistatic interactions between divergent genetic factors, as each lineage accumulated substitutions independently in their evolutionary history. Such negative genetic interactions can manifest as gene misexpression due to divergence in gene regulatory controls from mutations in cis-regulatory elements and trans-acting factors. Gene misexpression due to differences in regulatory controls can ultimately contribute to incompatibility within hybrids through developmental defects such as sterility and inviability. We sought to quantify the contributions of regulatory divergence to postzygotic reproductive isolation using sterile interspecies hybrids of two Caenorhabditis nematodes: Caenorhabditis briggsae and Caenorhabditis nigoni. We analyzed previous transcriptome profiles for two introgression lines with distinct homozygous X-linked fragments from C. briggsae in a C. nigoni genomic background that confers male sterility, owing to defects in spermatogenesis (Li R, et al. 2016. Specific down-regulation of spermatogenesis genes targeted by 22G RNAs in hybrid sterile males associated with an X-chromosome introgression. Genome Res. 26:1219-1232). Our analysis identified hundreds of genes that show distinct classes of nonadditive expression inheritance and regulatory divergence. We find that these nonoverlapping introgressions affect many of the same genes in the same way and demonstrate that the preponderance of transgressive gene expression is due to regulatory divergence involving compensatory and joint effects of cis- and trans-acting factors. The similar transcriptomic responses to nonoverlapping genetic perturbations of the X-chromosome implicate multiway incompatibilities as an important feature contributing to hybrid male sterility in this system.

Synthetic gene drives as an anthropogenic evolutionary force.

Genetic drive represents a fundamental evolutionary force that can exact profound change to the genetic composition of populations by biasing allele transmission. Herein I propose that the use of synthetic homing gene drives, the human-mediated analog of endogenous genetic drives, warrants the designation of 'genetic welding' as an anthropogenic evolutionary force. Conceptually, this distinction parallels that of artificial and natural selection. Genetic welding is capable of imposing complex and rapid heritable phenotypic change on entire populations, whether motivated by biodiversity conservation or public health. Unanticipated possible long-term evolutionary outcomes, however, demand further investigation and bioethical consideration. The emerging importance of genetic welding also compels our explicit recognition of genetic drive as an addition to the other four fundamental forces of evolution.

Temperature-dependent Small RNA Expression Depends on Wild Genetic Backgrounds of Caenorhabditis briggsae.

Geographically distinct populations can adapt to the temperature conditions of their local environment, leading to temperature-dependent fitness differences between populations. Consistent with local adaptation, phylogeographically distinct Caenorhabditis briggsae nematodes show distinct fitness responses to temperature. The genetic mechanisms underlying local adaptation, however, remain unresolved. To investigate the potential role of small noncoding RNAs in genotype-specific responses to temperature, we quantified small RNA expression using high-throughput sequencing of C. briggsae nematodes from tropical and temperate strain genotypes reared under three temperature conditions (14 °C, 20 °C, and 30 C). Strains representing both tropical and temperate regions showed significantly lower expression of PIWI-interacting RNAs (piRNAs) at high temperatures, primarily mapping to a large ∼7 Mb long piRNA cluster on chromosome IV. We also documented decreased expression of 22G-RNAs antisense to protein-coding genes and other genomic features at high rearing temperatures for the thermally-intolerant temperate strain genotype, but not for the tropical strain genotype. Reduced 22G-RNA expression was widespread along chromosomes and among feature types, indicative of a genome-wide response. Targets of the EGO-1/CSR-1 22G-RNA pathway were most strongly impacted compared with other 22G-RNA pathways, implicating the CSR-1 Argonaute and its RNA-dependent RNA polymerase EGO-1 in the genotype-dependent modulation of C. briggsae 22G-RNAs under chronic thermal stress. Our work suggests that gene regulation via small RNAs may be an important contributor to the evolution of local adaptations.

The distribution of mutational effects on fitness in Caenorhabditis elegans inferred from standing genetic variation.

The distribution of fitness effects (DFE) for new mutations is one of the most theoretically important but difficult to estimate properties in population genetics. A crucial challenge to inferring the DFE from natural genetic variation is the sensitivity of the site frequency spectrum to factors like population size change, population substructure, genome structure, and nonrandom mating. Although inference methods aim to control for population size changes, the influence of nonrandom mating remains incompletely understood, despite being a common feature of many species. We report the DFE estimated from 326 genomes of Caenorhabditis elegans, a nematode roundworm with a high rate of self-fertilization. We evaluate the robustness of DFE inferences using simulated data that mimics the genomic structure and reproductive life history of C. elegans. Our observations demonstrate how the combined influence of self-fertilization, genome structure, and natural selection on linked sites can conspire to compromise estimates of the DFE from extant polymorphisms with existing methods. These factors together tend to bias inferences toward weakly deleterious mutations, making it challenging to have full confidence in the inferred DFE of new mutations as deduced from standing genetic variation in species like C. elegans. Improved methods for inferring the DFE are needed to appropriately handle strong linked selection and selfing. These results highlight the importance of understanding the combined effects of processes that can bias our interpretations of evolution in natural populations.

Widespread misregulation of inter-species hybrid transcriptomes due to sex-specific and sex-chromosome regulatory evolution.

When gene regulatory networks diverge between species, their dysfunctional expression in inter-species hybrid individuals can create genetic incompatibilities that generate the developmental defects responsible for intrinsic post-zygotic reproductive isolation. Both cis- and trans-acting regulatory divergence can be hastened by directional selection through adaptation, sexual selection, and inter-sexual conflict, in addition to cryptic evolution under stabilizing selection. Dysfunctional sex-biased gene expression, in particular, may provide an important source of sexually-dimorphic genetic incompatibilities. Here, we characterize and compare male and female/hermaphrodite transcriptome profiles for sibling nematode species Caenorhabditis briggsae and C. nigoni, along with allele-specific expression in their F1 hybrids, to deconvolve features of expression divergence and regulatory dysfunction. Despite evidence of widespread stabilizing selection on gene expression, misexpression of sex-biased genes pervades F1 hybrids of both sexes. This finding implicates greater fragility of male genetic networks to produce dysfunctional organismal phenotypes. Spermatogenesis genes are especially prone to high divergence in both expression and coding sequences, consistent with a "faster male" model for Haldane's rule and elevated sterility of hybrid males. Moreover, underdominant expression pervades male-biased genes compared to female-biased and sex-neutral genes and an excess of cis-trans compensatory regulatory divergence for X-linked genes underscores a "large-X effect" for hybrid male expression dysfunction. Extensive regulatory divergence in sex determination pathway genes likely contributes to demasculinization of XX hybrids. The evolution of genetic incompatibilities due to regulatory versus coding sequence divergence, however, are expected to arise in an uncorrelated fashion. This study identifies important differences between the sexes in how regulatory networks diverge to contribute to sex-biases in how genetic incompatibilities manifest during the speciation process.

Speciation and the developmental alarm clock.

New species arise as the genomes of populations diverge. The developmental 'alarm clock' of speciation sounds off when sufficient divergence in genetic control of development leads hybrid individuals to infertility or inviability, the world awoken to the dawn of new species with intrinsic post-zygotic reproductive isolation. Some developmental stages will be more prone to hybrid dysfunction due to how molecular evolution interacts with the ontogenetic timing of gene expression. Considering the ontogeny of hybrid incompatibilities provides a profitable connection between 'evo-devo' and speciation genetics to better link macroevolutionary pattern, microevolutionary process, and molecular mechanisms. Here, we explore speciation alongside development, emphasizing their mutual dependence on genetic network features, fitness landscapes, and developmental system drift. We assess models for how ontogenetic timing of reproductive isolation can be predictable. Experiments and theory within this synthetic perspective can help identify new rules of speciation as well as rules in the molecular evolution of development.

Males, Outcrossing, and Sexual Selection in Caenorhabditis Nematodes.

Males of Caenorhabditis elegans provide a crucial practical tool in the laboratory, but, as the rarer and more finicky sex, have not enjoyed the same depth of research attention as hermaphrodites. Males, however, have attracted the attention of evolutionary biologists who are exploiting the C. elegans system to test longstanding hypotheses about sexual selection, sexual conflict, transitions in reproductive mode, and genome evolution, as well as to make new discoveries about Caenorhabditis organismal biology. Here, we review the evolutionary concepts and data informed by study of males of C. elegans and other Caenorhabditis We give special attention to the important role of sperm cells as a mediator of inter-male competition and male-female conflict that has led to drastic trait divergence across species, despite exceptional phenotypic conservation in many other morphological features. We discuss the evolutionary forces important in the origins of reproductive mode transitions from males being common (gonochorism: females and males) to rare (androdioecy: hermaphrodites and males) and the factors that modulate male frequency in extant androdioecious populations, including the potential influence of selective interference, host-pathogen coevolution, and mutation accumulation. Further, we summarize the consequences of males being common vs rare for adaptation and for trait divergence, trait degradation, and trait dimorphism between the sexes, as well as for molecular evolution of the genome, at both micro-evolutionary and macro-evolutionary timescales. We conclude that C. elegans male biology remains underexploited and that future studies leveraging its extensive experimental resources are poised to discover novel biology and to inform profound questions about animal function and evolution.

Molecular evolution across developmental time reveals rapid divergence in early embryogenesis.

Ontogenetic development hinges on the changes in gene expression in time and space within an organism, suggesting that the demands of ontogenetic growth can impose or reveal predictable pattern in the molecular evolution of genes expressed dynamically across development. Here, we characterize coexpression modules of the Caenorhabditis elegans transcriptome, using a time series of 30 points from early embryo to adult. By capturing the functional form of expression profiles with quantitative metrics, we find fastest evolution in the distinctive set of genes with transcript abundance that declines through development from a peak in young embryos. These genes are highly enriched for oogenic function and transient early zygotic expression, are nonrandomly distributed in the genome, and correspond to a life stage especially prone to inviability in interspecies hybrids. These observations conflict with the "early conservation model" for the evolution of development, although expression-weighted sequence divergence analysis provides some support for the "hourglass model." Genes in coexpression modules that peak toward adulthood also evolve fast, being hyper-enriched for roles in spermatogenesis, implicating a history of sexual selection and relaxation of selection on sperm as key factors driving rapid change to ontogenetically distinguishable coexpression modules of genes. We propose that these predictable trends of molecular evolution for dynamically expressed genes across ontogeny predispose particular life stages, early embryogenesis in particular, to hybrid dysfunction in the speciation process.

Genome structure predicts modular transcriptome responses to genetic and environmental conditions.

Understanding the plasticity, robustness and modularity of transcriptome expression to genetic and environmental conditions is crucial to deciphering how organisms adapt in nature. To test how genome architecture influences transcriptome profiles, we quantified expression responses for distinct temperature-adapted genotypes of the nematode Caenorhabditis briggsae when exposed to chronic temperature stresses throughout development. We found that 56% of the 8,795 differentially expressed genes show genotype-specific changes in expression in response to temperature (genotype-by-environment interactions, GxE). Most genotype-specific responses occur under heat stress, indicating that cold vs. heat stress responses involve distinct genomic architectures. The 22 co-expression modules that we identified differ in their enrichment of genes with genetic vs. environmental vs. interaction effects, as well as their genomic spatial distributions, functional attributes and rates of molecular evolution at the sequence level. Genes in modules enriched for simple effects of either genotype or temperature alone tend to evolve especially rapidly, consistent with disproportionate influence of adaptation or weaker constraint on these subsets of loci. Chromosome-scale heterogeneity in nucleotide polymorphism, however, rather than the scale of individual genes predominates as the source of genetic differences among expression profiles, and natural selection regimes are largely decoupled between coding sequences and noncoding flanking sequences that contain cis-regulatory elements. These results illustrate how the form of transcriptome modularity and genome structure contribute to predictable profiles of evolutionary change.

Reproductive transitions in plants and animals: selfing syndrome, sexual selection and speciation.

The evolution of predominant self-fertilisation frequently coincides with the evolution of a collection of phenotypes that comprise the 'selfing syndrome', in both plants and animals. Genomic features also display a selfing syndrome. Selfing syndrome traits often involve changes to male and female reproductive characters that were subject to sexual selection and sexual conflict in the obligatorily outcrossing ancestor, including the gametic phase for both plants and animals. Rapid evolution of reproductive traits, due to both relaxed selection and directional selection under the new status of predominant selfing, lays the genetic groundwork for reproductive isolation. Consequently, shifts in sexual selection pressures coupled to transitions to selfing provide a powerful paradigm for investigating the speciation process. Plant and animal studies, however, emphasise distinct selective forces influencing reproductive-mode transitions: genetic transmission advantage to selfing or reproductive assurance outweighing the costs of inbreeding depression vs the costs of males and meiosis. Here, I synthesise links between sexual selection, evolution of selfing and speciation, with particular focus on identifying commonalities and differences between plant and animal systems and pointing to areas warranting further synergy.

Neuro-genetic plasticity of Caenorhabditis elegans behavioral thermal tolerance.

BACKGROUND: Animal responses to thermal stimuli involve intricate contributions of genetics, neurobiology and physiology, with temperature variation providing a pervasive environmental factor for natural selection. Thermal behavior thus exemplifies a dynamic trait that requires non-trivial phenotypic summaries to appropriately capture the trait in response to a changing environment. To characterize the deterministic and plastic components of thermal responses, we developed a novel micro-droplet assay of nematode behavior that permits information-dense summaries of dynamic behavioral phenotypes as reaction norms in response to increasing temperature (thermal tolerance curves, TTC). RESULTS: We found that C. elegans TTCs shift predictably with rearing conditions and developmental stage, with significant differences between distinct wildtype genetic backgrounds. Moreover, after screening TTCs for 58 C. elegans genetic mutant strains, we determined that genes affecting thermosensation, including cmk-1 and tax-4, potentially play important roles in the behavioral control of locomotion at high temperature, implicating neural decision-making in TTC shape rather than just generalized physiological limits. However, expression of the transient receptor potential ion channel TRPA-1 in the nervous system is not sufficient to rescue rearing-dependent plasticity in TTCs conferred by normal expression of this gene, indicating instead a role for intestinal signaling involving TRPA-1 in the adaptive plasticity of thermal performance. CONCLUSIONS: These results implicate nervous system and non-nervous system contributions to behavior, in addition to basic cellular physiology, as key mediators of evolutionary responses to selection from temperature variation in nature.

Genetically Distinct Behavioral Modules Underlie Natural Variation in Thermal Performance Curves.

Thermal reaction norms pervade organismal traits as stereotyped responses to temperature, a fundamental environmental input into sensory and physiological systems. Locomotory behavior represents an especially plastic read-out of animal response, with its dynamic dependence on environmental stimuli presenting a challenge for analysis and for understanding the genomic architecture of heritable variation. Here we characterize behavioral reaction norms as thermal performance curves for the nematode Caenorhabditis briggsae, using a collection of 23 wild isolate genotypes and 153 recombinant inbred lines to quantify the extent of genetic and plastic variation in locomotory behavior to temperature changes. By reducing the dimensionality of the multivariate phenotypic response with a function-valued trait framework, we identified genetically distinct behavioral modules that contribute to the heritable variation in the emergent overall behavioral thermal performance curve. Quantitative trait locus mapping isolated regions on Chromosome II associated with locomotory activity at benign temperatures and Chromosome V loci related to distinct aspects of sensitivity to high temperatures, with each quantitative trait locus explaining up to 28% of trait variation. These findings highlight how behavioral responses to environmental inputs as thermal reaction norms can evolve through independent changes to genetically distinct modular components of such complex phenotypes.

Demographic consequences of reproductive interference in multi-species communities.

BACKGROUND: Reproductive interference can mediate interference competition between species through sexual interactions that reduce the fitness of one species by another. Theory shows that the positive frequency-dependent effects of such costly errors in mate recognition can dictate species coexistence or exclusion even with countervailing resource competition differences between species. While usually framed in terms of pre-mating or post-zygotic costs, reproductive interference manifests between individual Caenorhabditis nematodes from negative interspecies gametic interactions: sperm cells from interspecies matings can migrate ectopically to induce female sterility and premature death. The potential for reproductive interference to exert population level effects on Caenorhabditis trait evolution and community structure, however, remains unknown. RESULTS: Here we test whether a species that is superior in individual-level reproductive interference (C. nigoni) can exact negative demographic effects on competitor species that are superior in resource competition (C. briggsae and C. elegans). We observe coexistence over six generations and find evidence of demographic reproductive interference even under conditions unfavorable to its influence. C. briggsae and C. elegans show distinct patterns of reproductive interference in competitive interactions with C. nigoni. CONCLUSIONS: These results affirm that individual level negative effects of reproductive interference mediated by gamete interactions can ramify to population demography, with the potential to influence patterns of species coexistence separately from the effects of direct resource competition.

Genetic Contributions to Ectopic Sperm Cell Migration in Caenorhabditis Nematodes.

Reproductive barriers involving gametic incompatibilities can act to enhance population divergence and promote the persistence of species boundaries. Observing gametic interactions in internal fertilizing organisms, however, presents a considerable practical challenge to characterizing mechanisms of such gametic isolation. Here we exploit the transparency of Caenorhabditis nematodes to investigate gametic isolation mediated by sperm that can migrate to ectopic locations, with this sperm invasion capable of inducing female sterility and premature death. As a step toward identifying genetic factors and mechanisms associated with female susceptibility to sperm invasion, we characterized a panel of 25 C. elegans genetic mutants to test for effects on the incidence and severity of sperm invasion in both conspecific and inter-species matings. We found genetic perturbations to contribute to distinct patterns of susceptibility that identify ovulation dynamics and sperm guidance cues as modulators of ectopic sperm migration incidence and severity. Genotypes confer distinctive phenotypic sensitivities to the sperm from conspecific C. elegans males vs. heterospecific C. nigoni males, implicating evolution of functional divergence in the history of these species for components of sperm-reproductive tract interactions. Sexually-antagonistic co-evolution within species that drives divergent trait and molecular evolution between species provides a working model to explain mismatched species-specific gametic interactions that promote or mitigate ectopic sperm migration.

Genetic basis to hybrid inviability is more complex than hybrid male sterility in Caenorhabditis nematodes.

Hybrid male sterility often evolves before female sterility or inviability of hybrids, implying that the accumulation of divergence between separated lineages should lead hybrid male sterility to have a more polygenic basis. However, experimental evidence is mixed. Here, we use the nematodes Caenorhabditis remanei and C. latens to characterize the underlying genetic basis of asymmetric hybrid male sterility and hybrid inviability. We demonstrate that hybrid male sterility is consistent with a simple genetic basis, involving a single X-autosome incompatibility. We also show that hybrid inviability involves more genomic compartments, involving diverse nuclear-nuclear incompatibilities, a mito-nuclear incompatibility, and maternal effects. These findings demonstrate that male sensitivity to genetic perturbation may be genetically simple compared to hybrid inviability in Caenorhabditis and motivates tests of generality for the genetic architecture of hybrid incompatibility across the breadth of phylogeny.