Stanford Advisors

2023-24 Courses

All Publications

  • Evolution of insect innate immunity through domestication of bacterial toxins. Proceedings of the National Academy of Sciences of the United States of America Verster, K. I., Cinege, G., Lipinszki, Z., Magyar, L. B., Kurucz, E., Tarnopol, R. L., Abraham, E., Darula, Z., Karageorgi, M., Tamsil, J. A., Akalu, S. M., Ando, I., Whiteman, N. K. 2023; 120 (16): e2218334120


    Toxin cargo genes are often horizontally transferred by phages between bacterial species and are known to play an important role in the evolution of bacterial pathogenesis. Here, we show how these same genes have been horizontally transferred from phage or bacteria to animals and have resulted in novel adaptations. We discovered that two widespread bacterial genes encoding toxins of animal cells, cytolethal distending toxin subunit B (cdtB) and apoptosis-inducing protein of 56 kDa (aip56), were captured by insect genomes through horizontal gene transfer from bacteria or phages. To study the function of these genes in insects, we focused on Drosophila ananassae as a model. In the D. ananassae subgroup species, cdtB and aip56 are present as singular (cdtB) or fused copies (cdtB::aip56) on the second chromosome. We found that cdtB and aip56 genes and encoded proteins were expressed by immune cells, some proteins were localized to the wasp embryo's serosa, and their expression increased following parasitoid wasp infection. Species of the ananassae subgroup are highly resistant to parasitoid wasps, and we observed that D. ananassae lines carrying null mutations in cdtB and aip56 toxin genes were more susceptible to parasitoids than the wild type. We conclude that toxin cargo genes were captured by these insects millions of years ago and integrated as novel modules into their innate immune system. These modules now represent components of a heretofore undescribed defense response and are important for resistance to parasitoid wasps. Phage or bacterially derived eukaryotic toxin genes serve as macromutations that can spur the instantaneous evolution of novelty in animals.

    View details for DOI 10.1073/pnas.2218334120

    View details for PubMedID 37036995

  • Horizontal transfer of bacterial cytolethal distending toxin B genes to insects. Molecular biology and evolution Verster, K. I., Wisecaver, J. H., Karageorgi, M., Duncan, R. P., Gloss, A. D., Armstrong, E., Price, D. K., Menon, A. R., Ali, Z. M., Whiteman, N. K. 2019


    Horizontal gene transfer (HGT) events have played a major role in the evolution of microbial species, but their importance in animals is less clear. Here we report HGT of cytolethal distending toxin B (cdtB), prokaryotic genes encoding eukaryote-targeting DNase I toxins, into the genomes of vinegar flies (Diptera: Drosophilidae) and aphids (Hemiptera: Aphididae). We found insect-encoded cdtB genes are most closely related to orthologs from bacteriophage that infect Candidatus Hamiltonella defensa, a bacterial mutualistic symbiont of aphids that may confer resistance to parasitoid wasps. In drosophilids, cdtB orthologs are highly expressed during the parasitoid-prone larval stage and encode a protein with ancestral DNase activity. We show that cdtB has been domesticated by diverse insects and hypothesize that it functions in defense against their natural enemies.

    View details for DOI 10.1093/molbev/msz146

    View details for PubMedID 31236589

  • Evolution of chemosensory and detoxification gene families across herbivorous Drosophilidae. G3 (Bethesda, Md.) Peláez, J. N., Gloss, A. D., Goldman-Huertas, B., Kim, B., Lapoint, R. T., Pimentel-Solorio, G., Verster, K. I., Aguilar, J. M., Nelson Dittrich, A. C., Singhal, M., Suzuki, H. C., Matsunaga, T., Armstrong, E. E., Charboneau, J. L., Groen, S. C., Hembry, D. H., Ochoa, C. J., O'Connor, T. K., Prost, S., Zaaijer, S., Nabity, P. D., Wang, J., Rodas, E., Liang, I., Whiteman, N. K. 2023


    Herbivorous insects are exceptionally diverse, accounting for a quarter of all known eukaryotic species, but the genomic basis of adaptations that enabled this dietary transition remains poorly understood. Many studies have suggested that expansions and contractions of chemosensory and detoxification gene families - genes directly mediating interactions with plant chemical defenses - underlie successful plant colonization. However, this hypothesis has been challenging to test because the origins of herbivory in many insect lineages are ancient (>150 million years ago [mya]), obscuring genomic evolutionary patterns. Here, we characterized chemosensory and detoxification gene family evolution across Scaptomyza, a genus nested within Drosophila that includes a recently derived (<15 mya) herbivore lineage of mustard (Brassicales) specialists and carnation (Caryophyllaceae) specialists, and several non-herbivorous species. Comparative genomic analyses revealed that herbivorous Scaptomyza have among the smallest chemosensory and detoxification gene repertoires across 12 drosophilid species surveyed. Rates of gene turnover averaged across the herbivore clade were significantly higher than background rates in over half of the surveyed gene families. However, gene turnover was more limited along the ancestral herbivore branch, with only gustatory receptors and odorant binding proteins experiencing strong losses. The genes most significantly impacted by gene loss, duplication, or changes in selective constraint were those involved in detecting compounds associated with feeding on living plants (bitter or electrophilic phytotoxins) or their ancestral diet (fermenting plant volatiles). These results provide insight into the molecular and evolutionary mechanisms of plant-feeding adaptations and highlight gene candidates that have also been linked to other dietary transitions in Drosophila.

    View details for DOI 10.1093/g3journal/jkad133

    View details for PubMedID 37317982

  • Evolution of Olfactory Receptors Tuned to Mustard Oils in Herbivorous Drosophilidae MOLECULAR BIOLOGY AND EVOLUTION Matsunaga, T., Reisenman, C. E., Goldman-Huertas, B., Brand, P., Miao, K., Suzuki, H. C., Verster, K., Ramirez, S. R., Whiteman, N. K. 2022; 39 (2)


    The diversity of herbivorous insects is attributed to their propensity to specialize on toxic plants. In an evolutionary twist, toxins betray the identity of their bearers when herbivores coopt them as cues for host-plant finding, but the evolutionary mechanisms underlying this phenomenon are poorly understood. We focused on Scaptomyza flava, an herbivorous drosophilid specialized on isothiocyanate (ITC)-producing (Brassicales) plants, and identified Or67b paralogs that were triplicated as mustard-specific herbivory evolved. Using in vivo heterologous systems for the expression of olfactory receptors, we found that S. flava Or67bs, but not the homologs from microbe-feeding relatives, responded selectively to ITCs, each paralog detecting different ITC subsets. Consistent with this, S. flava was attracted to ITCs, as was Drosophila melanogaster expressing S. flava Or67b3 in the homologous Or67b olfactory circuit. ITCs were likely coopted as olfactory attractants through gene duplication and functional specialization (neofunctionalization and subfunctionalization) in S. flava, a recently derived herbivore.

    View details for DOI 10.1093/molbev/msab362

    View details for Web of Science ID 000767848900007

    View details for PubMedID 34963012

    View details for PubMedCentralID PMC8826531

  • Horizontal Transfer of Microbial Toxin Genes to Gall Midge Genomes GENOME BIOLOGY AND EVOLUTION Verster, K., Tarnopol, R. L., Akalu, S. M., Whiteman, N. K. 2021; 13 (9)


    A growing body of evidence has underscored the role of horizontal gene transfer (HGT) in animal evolution. Previously, we discovered the horizontal transfer of the gene encoding the eukaryotic genotoxin cytolethal distending toxin B (cdtB) from the pea aphid Acyrthosiphon pisum secondary endosymbiont (APSE) phages to drosophilid and aphid nuclear genomes. Here, we report cdtB in the nuclear genome of the gall-forming "swede midge" Contarinia nasturtii (Diptera: Cecidomyiidae) via HGT. We searched all available gall midge genome sequences for evidence of APSE-to-insect HGT events and found five toxin genes (aip56, cdtB, lysozyme, rhs, and sltxB) transferred horizontally to cecidomyiid nuclear genomes. Surprisingly, phylogenetic analyses of HGT candidates indicated APSE phages were often not the ancestral donor lineage of the toxin gene to cecidomyiids. We used a phylogenetic signal statistic to test a transfer-by-proximity hypothesis for animal HGT, which suggested that microbe-to-insect HGT was more likely between taxa that share environments than those from different environments. Many of the toxins we found in midge genomes target eukaryotic cells, and catalytic residues important for toxin function are conserved in insect copies. This class of horizontally transferred, eukaryotic cell-targeting genes is potentially important in insect adaptation.

    View details for DOI 10.1093/gbe/evab202

    View details for Web of Science ID 000731090000018

    View details for PubMedID 34450656

    View details for PubMedCentralID PMC8455502

  • Genome editing retraces the evolution of toxin resistance in the monarch butterfly NATURE Karageorgi, M., Groen, S. C., Sumbul, F., Pelaez, J. N., Verster, K. I., Aguilar, J. M., Hastings, A. P., Bernstein, S. L., Matsunaga, T., Astourian, M., Guerra, G., Rico, F., Dobler, S., Agrawal, A. A., Whiteman, N. K. 2019; 574 (7778): 409-+


    Identifying the genetic mechanisms of adaptation requires the elucidation of links between the evolution of DNA sequence, phenotype, and fitness1. Convergent evolution can be used as a guide to identify candidate mutations that underlie adaptive traits2-4, and new genome editing technology is facilitating functional validation of these mutations in whole organisms1,5. We combined these approaches to study a classic case of convergence in insects from six orders, including the monarch butterfly (Danaus plexippus), that have independently evolved to colonize plants that produce cardiac glycoside toxins6-11. Many of these insects evolved parallel amino acid substitutions in the α-subunit (ATPα) of the sodium pump (Na+/K+-ATPase)7-11, the physiological target of cardiac glycosides12. Here we describe mutational paths involving three repeatedly changing amino acid sites (111, 119 and 122) in ATPα that are associated with cardiac glycoside specialization13,14. We then performed CRISPR-Cas9 base editing on the native Atpα gene in Drosophila melanogaster flies and retraced the mutational path taken across the monarch lineage11,15. We show in vivo, in vitro and in silico that the path conferred resistance and target-site insensitivity to cardiac glycosides16, culminating in triple mutant 'monarch flies' that were as insensitive to cardiac glycosides as monarch butterflies. 'Monarch flies' retained small amounts of cardiac glycosides through metamorphosis, a trait that has been optimized in monarch butterflies to deter predators17-19. The order in which the substitutions evolved was explained by amelioration of antagonistic pleiotropy through epistasis13,14,20-22. Our study illuminates how the monarch butterfly evolved resistance to a class of plant toxins, eventually becoming unpalatable, and changing the nature of species interactions within ecological communities2,6-11,15,17-19.

    View details for DOI 10.1038/s41586-019-1610-8

    View details for Web of Science ID 000490988300066

    View details for PubMedID 31578524

    View details for PubMedCentralID PMC7039281