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.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Fig. 1: Diversity of morphological plasticity among three onthophagine species.
Fig. 2: Nutrition-responsive differential gene expression as a function of nutrition across three onthophagine species.
Fig. 3: Distribution of differentially expressed genes following different patterns of response to nutrition.
Fig. 4: Comparative analyses of dsx function in the regulation of horn growth.
Fig. 5: Gene expression variation (MAD) across plasticity categories.

Data availability

All data are available through NCBI’s Short Read Archive (BioProject accession: PRJNA608082).


  1. 1.

    Beldade, P., Mateus, A. R. & Keller, R. Evolution and molecular mechanisms of adaptive developmental plasticity. Mol. Ecol. 20, 1347–1363 (2011).

    PubMed  Google Scholar 

  2. 2.

    West-Eberhard, M. J. Developmental Plasticity and Evolution (Oxford Univ. Press, 2003).

  3. 3.

    Koyama, T., Mendes, C. C. & Mirth, C. K. Mechanisms regulating nutrition-dependent developmental plasticity through organ-specific effects in insects. Front. Physiol. 4, 263 (2013).

    PubMed  PubMed Central  Google Scholar 

  4. 4.

    Gotoh, H. et al. Developmental link between sex and nutrition; doublesex regulates sex-specific mandible growth via juvenile hormone signaling in stag beetles. PLoS Genet. 10, e1004098 (2014).

    PubMed  PubMed Central  Google Scholar 

  5. 5.

    Koyama, T. & Mirth, C. K. Unravelling the diversity of mechanisms through which nutrition regulates body size in insects. Curr. Opin. Insect Sci. 25, 1–8 (2018).

    PubMed  Google Scholar 

  6. 6.

    Mirth, C. K. & Shingleton, A. W. Integrating body and organ size in Drosophila: recent advances and outstanding problems. Front. Endocrinol. 3, 49 (2012).

    Google Scholar 

  7. 7.

    Emlen, D. J., Warren, I. A., Johns, A., Dworkin, I. & Lavine, L. C. A mechanism of extreme growth and reliable signaling in sexually selected ornaments and weapons. Science 337, 860–864 (2012).

    CAS  PubMed  Google Scholar 

  8. 8.

    Casasa, S. & Moczek, A. P. Insulin signalling’s role in mediating tissue-specific nutritional plasticity and robustness in the horn-polyphenic beetle Onthophagus taurus. Proc. R. Soc. Lond. B 285, 20181631 (2018).

    CAS  Google Scholar 

  9. 9.

    Xu, H. J. et al. Two insulin receptors determine alternative wing morphs in planthoppers. Nature 519, 464–467 (2015).

    CAS  PubMed  Google Scholar 

  10. 10.

    Fawcett, M. M. et al. Manipulation of insulin signaling phenocopies evolution of a host-associated polyphenism. Nat. Commun. 9, 1699 (2018).

    PubMed  PubMed Central  Google Scholar 

  11. 11.

    Kijimoto, T., Moczek, A. P. & Andrews, J. Diversification of doublesex function underlies morph-, sex-, and species-specific development of beetle horns. Proc. Natl Acad. Sci. USA 109, 20526–20531 (2012).

    CAS  PubMed  Google Scholar 

  12. 12.

    Kijimoto, T. & Moczek, A. P. Hedgehog signaling enables nutrition-responsive inhibition of an alternative morph in a polyphenic beetle. Proc. Natl Acad. Sci. USA 113, 5982–5987 (2016).

    CAS  PubMed  Google Scholar 

  13. 13.

    Snell-Rood, E. C. et al. Developmental decoupling of alternative phenotypes: insights from the transcriptomes of horn-polyphenic beetles. Evolution 65, 231–245 (2011).

    PubMed  Google Scholar 

  14. 14.

    Kijimoto, T. et al. The nutritionally responsive transcriptome of the polyphenic beetle Onthophagus taurus and the importance of sexual dimorphism and body region. Proc. R. Soc. Lond. B 281, 20142084 (2014).

    Google Scholar 

  15. 15.

    Klasberg, S., Bitard-Feildel, T. & Mallet, L. Computational identification of novel genes: current and future perspectives. Bioinform. Biol. Insights 10, 121–131 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Santos, M. E., Le Bouquin, A., Crumière, A. J. J. & Khila, A. Taxon-restricted genes at the origin of a novel trait allowing access to a new environment. Science 358, 386–390 (2017).

    CAS  PubMed  Google Scholar 

  17. 17.

    Pfennig, D. W. et al. Phenotypic plasticity’s impacts on diversification and speciation. Trends Ecol. Evol. 25, 459–467 (2010).

    PubMed  Google Scholar 

  18. 18.

    Moczek, A. P. et al. The role of developmental plasticity in evolutionary innovation. Proc. R. Soc. Lond. B 278, 2705–2713 (2011).

    Google Scholar 

  19. 19.

    Wund, M. A. Assessing the impacts of phenotypic plasticity on evolution. Integr. Comp. Biol. 52, 5–15 (2012).

    PubMed  Google Scholar 

  20. 20.

    Levis, N. A. & Pfennig, D. W. Evaluating ‘plasticity-first’ evolution in nature: key criteria and empirical approaches. Trends Ecol. Evol. 31, 563e574 (2016).

    Google Scholar 

  21. 21.

    Suzuki, Y. & Nijhout, H. F. Evolution of a polyphenism by genetic accommodation. Science 311, 650–652 (2006).

    CAS  PubMed  Google Scholar 

  22. 22.

    Ledón-Rettig, C. C., Pfennig, D. W. & Nascone-Yoder, N. Ancestral variation and the potential for genetic accommodation in larval amphibians: implications for the evolution of novel feeding strategies. Evol. Dev. 10, 316–325 (2008).

    PubMed  Google Scholar 

  23. 23.

    Scoville, A. G. & Pfrender, M. E. Phenotypic plasticity facilitates recurrent rapid adaptation to introduced predators. Proc. Natl Acad. Sci. USA 107, 4260–4263 (2010).

    CAS  PubMed  Google Scholar 

  24. 24.

    Sikkink, K. L., Reynolds, R. M., Ituarte, C. M., Cresko, W. & Phillips, P. C. Rapid evolution of phenotypic plasticity and shifting thresholds of genetic assimilation in the nematode Caenorhabditis remanei. G3 4, 1103–1112 (2014).

    CAS  PubMed  Google Scholar 

  25. 25.

    Susoy, V., Ragsdale, E. J., Kanzaki, N. & Sommer, R. J. Rapid diversification associated with a macroevolutionary pulse of developmental plasticity. eLife 4, e05463 (2015).

    PubMed Central  Google Scholar 

  26. 26.

    Walworth, N. G., Lee, M. D., Fu, F.-X., Hutchins, D. A. & Webb, E. A. Molecular and physiological evidence of genetic assimilation to high CO2 in the marine nitrogen fixer Trichodesmium. Proc. Natl Acad. Sci. USA 113, E7367–E7374 (2016).

    CAS  PubMed  Google Scholar 

  27. 27.

    Badyaev, A. V., Potticary, A. L. & Morrison, E. S. Most colorful example of genetic assimilation? Exploring the evolutionary destiny of recurrent phenotypic accommodation. Am. Nat. 190, 266–280 (2017).

    PubMed  Google Scholar 

  28. 28.

    Kulkarni, S. S., Denver, R. J., Gomez-Mestre, I. & Buchholz, D. R. Genetic accommodation via modified endocrine signalling explains phenotypic divergence among spadefoot toad species. Nat. Commun. 8, 993 (2017).

    PubMed  PubMed Central  Google Scholar 

  29. 29.

    Levis, N. A., Isdaner, A. J. & Pfennig, D. W. Morphological novelty emerges from pre-existing phenotypic plasticity. Nat. Ecol. Evol. 2, 1289–1297 (2018).

    PubMed  Google Scholar 

  30. 30.

    Levis, N. A., Serrato-Capuchina, A. & Pfennig, D. W. Genetic accommodation in the wild: evolution of gene expression plasticity during character displacement. J. Evol. Biol. 30, 1712–1723 (2017).

    CAS  PubMed  Google Scholar 

  31. 31.

    Renn, S. C. P. & Schummer, M. E. Genetic accommodation and behavioural evolution: insights from genomic studies. Anim. Behav. 85, 1012–1022 (2013).

    Google Scholar 

  32. 32.

    Génier, F. & Moretto, P. Digitonthophagus Balthasar, 1959: taxonomy, systematics, and morphological phylogeny of the genus revealing an African species complex (Coleoptera: Scarabaeidae: Scarabaeinae). Zootaxa 4248, 1–110 (2017).

    PubMed  Google Scholar 

  33. 33.

    Emlen, D. J., Hunt, J. & Simmons, L. W. Evolution of sexual dimorphism and male dimorphism in the expression of beetle horns: phylogenetic evidence for modularity, evolutionary lability, and constraint. Am. Nat. 166, S42–S68 (2005).

    PubMed  Google Scholar 

  34. 34.

    Nourmohammad, A. et al. Adaptive evolution of gene expression in Drosophila. Cell Rep. 20, 1385–1395 (2017).

    CAS  PubMed  Google Scholar 

  35. 35.

    Signor, S. A. & Nuzhdin, S. V. The evolution of gene expression in cis and trans. Trends Genet. 34, 532–544 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Zdobnov, E. M. et al. OrthoDB v9.1: cataloging evolutionary and functional annotations for animal, fungal, plant, archaeal, bacterial and viral orthologs. Nucleic Acids Res. 45, D744–D749 (2017).

    CAS  PubMed  Google Scholar 

  37. 37.

    Langfelder, P. & Horvath, S. WGCNA: an R package for weighted correlation network analysis. BMC Bioinform. 9, 559 (2008).

    Google Scholar 

  38. 38.

    Daniels, E. V., Murad, R., MortazaviA & Reed, R. D. Extensive transcriptional response associated with seasonal plasticity of butterfly wing patterns. Mol. Ecol. 23, 6123–6134 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Ghalambor, C. K. et al. Non-adaptive plasticity potentiates rapid adaptive evolution of gene expression in nature. Nature 525, 372–377 (2015).

    CAS  PubMed  Google Scholar 

  40. 40.

    Schrader, L., Helantera, H. & Oettler, J. Accelerated evolution of developmentally biased genes in the tetraphenic ant Cardiocondyla obscurior. Mol. Biol. Evol. 34, 535–544 (2016).

    PubMed Central  Google Scholar 

  41. 41.

    Kenkel, C. D. & Matz, M. V. Gene expression plasticity as a mechanism of coral adaptation to a variable environment. Nat. Ecol. Evol. 1, 0014 (2017).

    Google Scholar 

  42. 42.

    Alaux, C. et al. Honey bee aggression supports a link between gene regulation and behavioral evolution. Proc. Natl Acad. Sci. USA 106, 15400–15405 (2009).

    Google Scholar 

  43. 43.

    Moczek, A. P. & Nijhout, H. F. A method for sexing third instar larvae of the genus Onthophagus LATREILLE (Coleoptera: Scarabaeidae). Coleopt. Bull. 56, 279–284 (2002).

    Google Scholar 

  44. 44.

    Ledón-Rettig, C. C., Zattara, E. E. & Moczek, A. P. Asymmetric interactions between doublesex and sex- and tissue-specific target genes mediate sexual dimorphism in beetles. Nat. Commun. 8, 14593 (2017).

    PubMed  PubMed Central  Google Scholar 

  45. 45.

    Bolger, A. M., Lohse, M. & Usadel, B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30, 2114–2120 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. 46.

    Haas, B. J. et al. De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nat. Protoc. 8, 1494–1512 (2013).

    CAS  PubMed  Google Scholar 

  47. 47.

    Bryant, D. M. et al. A tissue-mapped axolotl de novo transcriptome enables identification of limb regeneration factors. Cell Rep. 18, 762–776 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Zattara, E. E., Hughes, D. S. T., Richards, S., Kijimoto, T. & Mocze, A. P. Onthophagus taurus genome annotations v0.5.3. Ag Data Commons (2016).

  49. 49.

    McCarthy, J. D., Chen, Y. & Smyth, K. G. Differential expression analysis of multifactor RNA-seq experiments with respect to biological variation. Nucleic Acids Res. 40, 4288–4297 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  50. 50.

    Love, M. I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550 (2014).

    PubMed  PubMed Central  Google Scholar 

  51. 51.

    R Development Core Team R: A Language and Environment for Statistical Computing (R Foundation for Statistical Computing, 2011);

  52. 52.

    Meyer, J. M. et al. Draft genome of the scarab beetle Oryctes borbonicus en La Réunion Island. Genome Biol. Evol. 8, 2093–2105 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  53. 53.

    Moczek, A. P. & Rose, D. J. Differential recruitment of limb patterning genes during development and diversification of beetle horns. Proc. Natl Acad. Sci. USA 106, 8992–899 (2009).

    CAS  PubMed  Google Scholar 

  54. 54.

    Moczek, A. P. A matter of measurements: challenges and approaches in the comparative analysis of static allometries. Am. Nat. 167, 606–611 (2006).

    PubMed  Google Scholar 

  55. 55.

    Snell-Rood, E. C. & Moczek, A. P. Insulin signaling as a mechanism underlying developmental plasticity: the role of FOXO in a nutritional polyphenism. PLoS ONE 7, e34857 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

Download references


We thank the Center for Genomics and Bioinformatics at Indiana University, which conducted sequencing and library construction, the Zdobnov laboratory for their guidance using OrthoDB and the National Center for Genome Analysis. We sincerely thank C. Ledón-Rettig, T. Ledón-Rettig, S. Close and W. Arnold for collecting beetles in the wild. This research was made possible through support by National Science Foundation grant nos. IOS 1256689 and 1901680 to A.P.M., as well as grant no. 61369 from the John Templeton Foundation. The opinions, interpretations, conclusions and recommendations are those of the authors and are not necessarily endorsed by the National Science Foundation or John Templeton Foundation.

Author information




S.C., E.E.Z. and A.P.M. designed the experiments. S.C. conducted the experiments (phenotyping, tissue dissection and RNA extraction). S.C., E.E.Z. and A.P.M. analysed and interpreted the data. S.C., E.E.Z. and A.P.M. wrote the manuscript.

Corresponding authors

Correspondence to Sofia Casasa or Eduardo E. Zattara.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Individuals used for dissections.

Relative pupal body size- horn size allometry for the three species used in this study (D. gazella: blue; O. taurus: red; O. sagittarius: yellow). Additionally, the novel, anterior head horn size of O. sagittarius is shown in white. Small and large individuals of each species that were used for dissections are shown in triangles. Relative body size was used to control for absolute body size differences across species and was calculated as the residual from the mean over the species’ body size range.

Supplementary information

Supplementary Information

Supplementary Tables 1–12, results and discussion, and methods.

Reporting Summary

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Casasa, S., Zattara, E.E. & Moczek, A.P. Nutrition-responsive gene expression and the developmental evolution of insect polyphenism. Nat Ecol Evol 4, 970–978 (2020).

Download citation

Further reading


Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing