Innexins facilitate cell-cell communication by forming gap junctions or non-junctional hemichannels, which play important roles in metabolic, chemical, ionic, and electrical coupling. The lack of knowledge regarding the evolution and role of these channels in ctenophores (comb jellies), the likely sister group to the rest of animals, represents a substantial gap in our understanding of the evolution of intercellular communication in animals. Here we identify and phylogenetically characterize the complete set of innexins of four ctenophores: Mnemiopsis leidyi, Hormiphora californensis, Pleurobrachia bachei, and Beroe ovata. Our phylogenetic analyses suggest that ctenophore innexins diversified independently from those of other animals and were established early in the emergence of ctenophores. We identified a four-innexin genomic cluster, which was present in the last common ancestor of these four species and has been largely maintained in these lineages. Evidence from correlated spatial and temporal gene expression of the M. leidyi innexin cluster suggest that this cluster has been maintained due to constraints related to gene regulation. We describe basic electrophysiological properties of putative ctenophore hemichannels from muscle cells using intracellular recording techniques, showing substantial overlap with the properties of bilaterian innexin channels. Together, our results suggest that the last common ancestor of animals had gap junctional channels also capable of forming functional innexin hemichannels, and that innexin genes have independently evolved in major lineages throughout Metazoa.

New mutations and standing genetic variations contribute significantly to repeated phenotypic evolution in sticklebacks. However, less is known about the role of introgression in this process. We analysed taxonomically and geographically comprehensive genomic data from Pungitius sticklebacks to decipher the extent of introgression and its consequences for the diversification of this genus. Our results demonstrate that introgression is more prevalent than suggested by earlier studies. Although gene flow was generally bidirectional, it was often asymmetric and left unequal genomic signatures in hybridizing species, which might, at least partly, be due to biased hybridization and/or population size differences. In several cases, introgression of variants from one species to another was accompanied by transitions of pelvic and/or lateral plate structures - important diagnostic traits in Pungitius systematics - and frequently left signatures of adaptation in the core gene regulatory networks of armor trait development. This finding suggests that introgression has been an important source of genetic variation and enabled phenotypic convergence among Pungitius sticklebacks. The results highlight the importance of introgression of genetic variation as a source of adaptive variation underlying key ecological and taxonomic traits. Taken together, our study indicates that introgression-driven convergence likely explains the longstanding challenges in resolving the taxonomy and systematics of this small but phenotypically highly diverse group of fish.

Freshwater Unionid bivalves have recently faced ecological upheaval through pollution, barriers to dispersal, harvesting, and changes in _sh-host prevalence. Currently, over 70% of species in North America are threatened, endangered or extinct. To characterize the genetic response to recent selective pressures, we collected population genetic data for one successful bivalve species, Megalonaias nervosa. We identify megabase sized regions that are nearly monomorphic across the population, signals of strong, recent selection reshaping diversity across 73Mb total. These signatures of selection are greater than is commonly seen in population genetic models. We observe 102 duplicate genes with high dN/dS on terminal branches among regions with sweeps, suggesting that gene duplication is a causative mechanism of recent adaptation in M. nervosa. Genes in sweeps reect functional classes important for Unionid survival, including anticoagulation genes important for _sh host parasitization, detox genes, mitochondria management, and shell formation. We identify sweeps in regions with no known functional impacts, suggesting mechanisms of adaptation that deserve greater attention in future work on species survival. In contrast, polymorphic transposable elements appear to be detrimental and underrepresented among regions with sweeps. TE site frequency spectra are skewed toward singleton variants, and TEs among regions with sweeps are present at low frequency. Our work suggests that duplicate genes are an essential source of genetic novelty that has helped this successful species succeed in environments where others have struggled. These results suggest that gene duplications deserve greater attention in non-model population genomics, especially in species that have recently faced sudden environmental challenges.

Bacterial evolution of antibiotic resistance frequently has deleterious side effects on microbial growth, virulence, and susceptibility to other antimicrobial agents. However, it is unclear how these trade-offs could be utilized for manipulating antibiotic resistance in the clinic, not least because the underlying molecular mechanisms are poorly understood. Using laboratory evolution, we demonstrate that clinically relevant resistance mutations in Escherichia coli constitutively rewire a large fraction of the transcriptome in a repeatable and stereotypic manner. Strikingly, lineages adapted to functionally distinct antibiotics and having no resistance mutations in common show a wide range of parallel gene expression changes that alter oxidative stress response, iron homeostasis, and the composition of the bacterial outer membrane and cell surface. These common physiological alterations are associated with changes in cell morphology and enhanced sensitivity to antimicrobial peptides. Finally, the constitutive transcriptomic changes induced by resistance mutations are largely distinct from those induced by antibiotic stresses in the wild type. This indicates a limited role for genetic assimilation of the induced antibiotic stress response during resistance evolution. Our work suggests that diverse resistance mutations converge on similar global transcriptomic states that shape genetic susceptibility to antimicrobial compounds.

Sunflowers of the genus Helianthus are models for hybridization research and contain three of the best-studied examples of homoploid hybrid speciation. To understand a broader picture of hybridization within the annual sunflowers, we used whole-genome resequencing to conduct a phylogenomic analysis and test for gene flow between lineages. We find that all annual sunflower species tested have evidence of admixture, suggesting hybridization was common during the radiation of the genus. Support for the major species tree decreases with increasing recombination rate, consistent with hybridization and introgression contributing to discordant topologies. Admixture graphs found hybridization to be associated with the origins of the three putative hybrid species (Helianthus anomalus, Helianthus deserticola, and Helianthus paradoxus). However, the hybridization events are more ancient than suggested by previous work. Furthermore, H. anomalus and H. deserticola appear to have arisen from a single hybridization event involving an unexpected donor, rather than through multiple independent events as previously proposed. This means our results are consistent with, but not definitive proof of, two ancient independent homoploid hybrid speciation events in the genus. Using a broader data set that covers the whole Helianthus genus, including perennial species, we find that signals of introgression span the genus and beyond, suggesting highly divergent introgression and/or the sorting of ancient haplotypes. Thus, Helianthus can be viewed as a syngameon in which largely reproductively isolated species are linked together by occasional or frequent gene flow.

Evolution of sequence-specific transcription factors clearly drives lineage-specific innovations, but less is known about how changes in the central transcriptional machinery may contribute to evolutionary transformations. In particular, transcriptional regulators are rich in intrinsically disordered regions that appear to be magnets for evolutionary innovation. The C-terminal Binding Protein (CtBP) is a transcriptional corepressor derived from an ancestral lineage of alpha hydroxyacid dehydrogenases; it is found in mammals and invertebrates, and features a core NAD-binding domain as well as an unstructured C-terminus (CTD) of unknown function. CtBP can act on promoters and enhancers to repress transcription through chromatin-linked mechanisms. Our comparative phylogenetic study shows that CtBP is a bilaterian innovation whose CTD of about 100 residues is present in almost all orthologs. CtBP CTDs contain conserved blocks of residues and retain a predicted disordered property, despite having variations in the primary sequence. Interestingly, the structure of the C-terminus has undergone radical transformation independently in certain lineages including flatworms and nematodes. Also contributing to CTD diversity is the production of myriad alternative RNA splicing products, including the production of "short" tailless forms of CtBP in Drosophila. Additional diversity stems from multiple gene duplications in vertebrates, where up to five CtBP orthologs have been observed. Vertebrate lineages show fewer major modifications in the unstructured CTD, possibly because gene regulatory constraints of the vertebrate body plan place specific constraints on this domain. Our study highlights the rich regulatory potential of this previously unstudied domain of a central transcriptional regulator.

Migratory birds play a critical role in the rapid spread of highly pathogenic avian influenza (HPAI) H5N8 virus clade 2.3.4.4 across Eurasia. Elucidating the timing and pattern of virus transmission is essential therefore for understanding the spatial dissemination of these viruses. In this study, we surveyed >27,000 wild birds in China, tracked the year-round migration patterns of 20 bird species across China since 2006, and generated new HPAI H5N8 virus genomic data. Using this new data set, we investigated the seasonal transmission dynamics of HPAI H5N8 viruses across Eurasia. We found that introductions of HPAI H5N8 viruses to different Eurasian regions were associated with the seasonal migration of wild birds. Moreover, we report a backflow of HPAI H5N8 virus lineages from Europe to Asia, suggesting that Europe acts as both a source and a sink in the global HPAI virus transmission network.

Single nucleotide polymorphisms are the most common type of genetic variation, but how these variants contribute to the adaptation of complex phenotypes is largely unknown. Experimental evolution and genome-wide association studies have demonstrated that variation in the PPARγ-homolog Eip75B has associated with longevity and life-history differences in the fruit fly Drosophila melanogaster. Using RNAi knockdown, we first demonstrate that reduced expression of Eip75B in adult flies affects lifespan, egg-laying rate, and egg volume. We then tested the effects of a naturally occurring SNP within a cis-regulatory domain of Eip75B by applying two complementary approaches: a Mendelian randomization approach using lines of the Drosophila Genetic Reference Panel, and allelic replacement using precise CRISPR/Cas9-induced genome editing. Our experiments reveal that this natural polymorphism has a significant pleiotropic effect on fecundity and egg-to-adult viability, but not on longevity or other life-history traits. Our results provide a rare functional validation at the nucleotide level and identify a natural allelic variant affecting fitness and life-history adaptation.

Island ecosystems provide natural laboratories to assess the impacts of isolation on population persistence. However, most studies of persistence have focused on a single species, without comparisons to other organisms they interact with in the ecosystem. The case study of moose and gray wolves on Isle Royale allows for a direct contrast of genetic variation in isolated populations that have experienced dramatically differing population trajectories over the past decade. Whereas the Isle Royale wolf population recently declined nearly to extinction due to severe inbreeding depression, the moose population has thrived and continues to persist, despite having low genetic diversity and being isolated for ∼120 years. Here, we examine the patterns of genomic variation underlying the continued persistence of the Isle Royale moose population. We document high levels of inbreeding in the population, roughly as high as the wolf population at the time of its decline. However, inbreeding in the moose population manifests in the form of intermediate-length runs of homozygosity suggestive of historical inbreeding and purging, contrasting with the long runs of homozygosity observed in the smaller wolf population. Using simulations, we confirm that substantial purging has likely occurred in the moose population. However, we also document notable increases in genetic load, which could eventually threaten population viability over the long term. Overall, our results demonstrate a complex relationship between inbreeding, genetic diversity, and population viability that highlights the use of genomic datasets and computational simulation tools for understanding the factors enabling persistence in isolated populations.

Studying the evolutionary history of gene families is a challenging and exciting task with a wide range of implications. In addition to exploring fundamental questions about the origin and evolution of genes, disentangling their evolution is also critical to those who do functional/structural studies to allow a deeper and more precise interpretation of their results in an evolutionary context. The sirtuin gene family is a group of genes that are involved in a variety of biological functions mostly related to aging. Their duplicative history is an open question, as well as the definition of the repertoire of sirtuin genes among vertebrates. Our results show a well-resolved phylogeny that represents an improvement in our understanding of the duplicative history of the sirtuin gene family. We identified a new sirtuin gene family member (SIRT3.2) that was apparently lost in the last common ancestor of amniotes but retained in all other groups of jawed vertebrates. According to our experimental analyses, elephant shark SIRT3.2 protein is located in mitochondria, the overexpression of which leads to an increase in cellular levels of ATP. Moreover, in vitro analysis demonstrated that it has deacetylase activity being modulated in a similar way to mammalian SIRT3. Our results indicate that there are at least eight sirtuin paralogs among vertebrates and that all of them can be traced back to the last common ancestor of the group that existed between 676 and 615 millions of years ago.

Asymmetries are essential for proper organization and function of organ systems. Genetic studies in bilaterians have shown signaling through the Nodal/Smad2 pathway plays a key, conserved role in the establishment of body asymmetries. Although the main molecular players in the network for the establishment of left-right asymmetry (LRA) have been deeply described in deuterostomes, little is known about the regulation of Nodal signaling in spiralians. Here, we identified orthologs of the egf-cfc gene, a master regulator of the Nodal pathway in vertebrates, in several invertebrate species, which includes the first evidence of its presence in non-deuterostomes. Our functional experiments indicate that despite being present, egf-cfc does not play a role in the establishment of LRA in gastropods. However, experiments in zebrafish suggest that a single amino acid mutation in the egf-cfc gene in at least the common ancestor of chordates was the necessary step to induce a gain of function in LRA regulation. This study shows that the egf-cfc gene likely appeared in the ancestors of deuterostomes and "protostomes", before being adopted as a mechanism to regulate the Nodal pathway and the establishment of LRA in some lineages of deuterostomes.

Some viruses (e.g., human immunodeficiency virus 1 and severe acute respiratory syndrome coronavirus 2) have been experimentally proposed to accelerate features of human aging and of cellular senescence. These observations, along with evolutionary considerations on viral fitness, raised the more general puzzling hypothesis that, beyond documented sources in human genetics, aging in our species may also depend on virally encoded interactions distorting our aging to the benefits of diverse viruses. Accordingly, we designed systematic network-based analyses of the human and viral protein interactomes, which unraveled dozens of viruses encoding proteins experimentally demonstrated to interact with proteins from pathways associated with human aging, including cellular senescence. We further corroborated our predictions that specific viruses interfere with human aging using published experimental evidence and transcriptomic data; identifying influenza A virus (subtype H1N1) as a major candidate age distorter, notably through manipulation of cellular senescence. By providing original evidence that viruses may convergently contribute to the evolution of numerous age-associated pathways through co-evolution, our network-based and bipartite network-based methodologies support an ecosystemic study of aging, also searching for genetic causes of aging outside a focal aging species. Our findings, predicting age distorters and targets for anti-aging therapies among human viruses, could have fundamental and practical implications for evolutionary biology, aging study, virology, medicine, and demography.

It has recently been proposed that lower mutation rates in gene bodies compared with upstream and downstream sequences in Arabidopsis thaliana are the result of an "adaptive" modification of the rate of beneficial and deleterious mutations in these functional regions. This claim was based both on analyses of mutation accumulation lines and on population genomics data. Here, we show that several questionable assumptions were used in the population genomics analyses. In particular, we demonstrate that the difference between gene bodies and less selectively constrained sequences in the magnitude of Tajima's D can in principle be explained by the presence of sites subject to purifying selection and does not require lower mutation rates in regions experiencing selective constraints.

Extreme environments, such as Antarctic habitats, present major challenges for many biological processes. Antarctic icefishes (Crynotothenioidea) represent a compelling system to investigate the molecular basis of adaptation to cold temperatures. Here we explore how the sub-zero habitats of Antarctic icefishes have impacted rhodopsin (RH1) function, the temperature-sensitive dim-light visual pigment found in rod photoreceptors. Using likelihood-models and ancestral reconstruction, we find that accelerated evolutionary rates in icefish RH1 underlie unique amino acid mutations absent from other deep-dwelling fishes, introduced before (S160A) and during (V259M) the onset of modern polar conditions. Functional assays reveal that these mutations red-shift rhodopsin spectral absorbance, consistent with spectral irradiance under sea-ice. These mutations also lower the activation energy associated with retinal release of the light-activated RH1, and accelerate its return to the dark-state, likely compensating for a cold-induced decrease in kinetic rates. These are adaptations in key properties of rhodopsin that mediate rod sensitivity and visual performance in the cold dark seas of the Antarctic.