Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Morphological evolution through multiple cis-regulatory mutations at a single gene


One central, and yet unsolved, question in evolutionary biology is the relationship between the genetic variants segregating within species and the causes of morphological differences between species. The classic neo-darwinian view postulates that species differences result from the accumulation of small-effect changes at multiple loci. However, many examples support the possible role of larger abrupt changes in the expression of developmental genes in morphological evolution1,2,3. Although this evidence might be considered a challenge to a neo-darwinian micromutationist view of evolution, there are currently few examples of the actual genes causing morphological differences between species4,5,6,7,8,9,10. Here we examine the genetic basis of a trichome pattern difference between Drosophila species, previously shown to result from the evolution of a single gene, shavenbaby (svb), probably through cis-regulatory changes6. We first identified three distinct svb enhancers from D. melanogaster driving reporter gene expression in partly overlapping patterns that together recapitulate endogenous svb expression. All three homologous enhancers from D. sechellia drive expression in modified patterns, in a direction consistent with the evolved svb expression pattern. To test the influence of these enhancers on the actual phenotypic difference, we conducted interspecific genetic mapping at a resolution sufficient to recover multiple intragenic recombinants. This functional analysis revealed that independent genetic regions upstream of svb that overlap the three identified enhancers are collectively required to generate the D. sechellia trichome pattern. Our results demonstrate that the accumulation of multiple small-effect changes at a single locus underlies the evolution of a morphological difference between species. These data support the view that alleles of large effect that distinguish species may sometimes reflect the accumulation of multiple mutations of small effect at select genes.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Modified svb expression underlies the evolved trichome pattern of D. sechellia.
Figure 2: Three enhancer regions, which collectively recapitulate the svb expression pattern, have evolved in D. sechellia.
Figure 3: High-resolution interspecific recombination mapping identifies three enhancer regions of svb that caused evolution of the D. sechellia trichome pattern.
Figure 4: A possible model of the evolutionary path of the D. sechellia trichome pattern.


  1. Carroll, S. B., Grenier, J. K. & Weatherbee, S. D. From DNA to Diversity: Molecular Genetics and the Evolution of Animal Design (Blackwell Science, Malden, 2001)

    Google Scholar 

  2. Marcellini, S. & Simpson, P. Two or four bristles: functional evolution of an enhancer of scute in Drosophilidae. PLoS Biol. 4, e386 (2006)

    Article  Google Scholar 

  3. Abzhanov, A., Protas, M., Grant, B. R., Grant, P. R. & Tabin, C. J. Bmp4 and morphological variation of beaks in Darwin’s finches. Science 305, 1462–1465 (2004)

    Article  ADS  CAS  Google Scholar 

  4. Wang, X. & Chamberlin, H. M. Multiple regulatory changes contribute to the evolution of the Caenorhabditis lin-48 ovo gene. Genes Dev. 16, 2345–2349 (2002)

    Article  CAS  Google Scholar 

  5. Doebley, J., Stec, A. & Gustus, C. teosinte branched1 and the origin of maize: Evidence for epistasis and the evolution of dominance. Genetics 141, 333–346 (1995)

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Sucena, E. & Stern, D. L. Divergence of larval morphology between Drosophila sechellia and its sibling species caused by cis-regulatory evolution of ovo/shaven-baby. Proc. Natl Acad. Sci. USA 97, 4530–4534 (2000)

    Article  ADS  CAS  Google Scholar 

  7. Stern, D. L. A role of Ultrabithorax in morphological differences between Drosophila species. Nature 396, 463–466 (1998)

    Article  ADS  CAS  Google Scholar 

  8. Yoon, H. S. & Baum, D. A. Transgenic study of parallelism in plant morphological evolution. Proc. Natl Acad. Sci. USA 101, 6524–6529 (2004)

    Article  ADS  CAS  Google Scholar 

  9. Wang, H. et al. The origin of the naked grains of maize. Nature 436, 714–719 (2005)

    Article  ADS  CAS  Google Scholar 

  10. Hay, A. & Tsiantis, M. The genetic basis for differences in leaf form between Arabidopsis thaliana and its wild relative Cardamine hirsuta. Nature Genet. 38, 942–947 (2006)

    Article  CAS  Google Scholar 

  11. Bokor, P. & DiNardo, S. The roles of hedgehog, wingless and lines in patterning the dorsal epidermis in Drosophila. Development 122, 1083–1092 (1996)

    CAS  PubMed  Google Scholar 

  12. Payre, F., Vincent, A. & Carreno, S. ovo/svb integrates Wingless and DER pathways to control epidermis differentiation. Nature 400, 271–275 (1999)

    Article  ADS  CAS  Google Scholar 

  13. Chanut-Delalande, H., Fernandes, I., Roch, F., Payre, F. & Plaza, S. Shavenbaby couples patterning to epidermal cell shape control. PLoS Biol. 4, e290 (2006)

    Article  Google Scholar 

  14. Sucena, E., Delon, I., Jones, I., Payre, F. & Stern, D. L. Regulatory evolution of shavenbaby/ovo underlies multiple cases of morphological parallelism. Nature 424, 935–938 (2003)

    Article  ADS  CAS  Google Scholar 

  15. Khila, A., El Haidani, A., Vincent, A., Payre, F. & Souda, S. I. The dual function of ovo/shavenbaby in germline and epidermis differentiation is conserved between Drosophila melanogaster and the olive fruit fly Bactrocera oleae. Insect Biochem. Mol. Biol. 33, 691–699 (2003)

    Article  CAS  Google Scholar 

  16. Mével-Ninio, M., Terracol, R., Salles, C., Vincent, A. & Payre, F. ovo, a Drosophila gene required for ovarian development, is specifically expressed in the germline and shares most of its coding sequences with shavenbaby, a gene involved in embryo patterning. Mech. Dev. 49, 83–95 (1995)

    Article  Google Scholar 

  17. Clark, R. M., Wagler, T. N., Quijada, P. & Doebley, J. A distant upstream enhancer at the maize domestication gene tb1 has pleiotropic effects on plant and inflorescent architecture. Nature Genet. 38, 594–597 (2006)

    Article  CAS  Google Scholar 

  18. True, J. R., Mercer, J. M. & Laurie, C. C. Differences in crossover frequency and distribution among three sibling species of Drosophila. Genetics 142, 507–523 (1996)

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Nusslein-Volhard, C. & Wieschaus, E. Mutations affecting segment number and polarity in Drosophila. Nature 287, 795–801 (1980)

    Article  ADS  CAS  Google Scholar 

  20. Wiellette, E. L. & McGinnis, W. Hox genes differentially regulate Serrate to generate segment-specific structures. Development 126, 1985–1995 (1999)

    CAS  PubMed  Google Scholar 

  21. Walters, J. W., Munoz, C., Paaby, A. B. & Dinardo, S. Serrate-Notch signaling defines the scope of the initial denticle field by modulating EGFR activation. Dev. Biol. 286, 415–426 (2005)

    Article  CAS  Google Scholar 

  22. Hatini, V., Green, R. B., Lengyel, J. A., Bray, S. J. & Dinardo, S. The Drumstick/Lines/Bowl regulatory pathway links antagonistic Hedgehog and Wingless signaling inputs to epidermal cell differentiation. Genes Dev. 19, 709–718 (2005)

    Article  CAS  Google Scholar 

  23. Delon, I., Chanut-Delalande, H. & Payre, F. The Ovo/Shavenbaby transcription factor specifies actin remodelling during epidermal differentiation in Drosophila. Mech. Dev. 120, 747–758 (2003)

    Article  CAS  Google Scholar 

  24. Stam, L. F. & Laurie, C. C. Molecular dissection of a major gene effect on a quantitative trait: The level of alcohol dehydrogenase expression in Drosophila melanogaster. Genetics 144, 1559–1564 (1996)

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Prud’homme, B. et al. Repeated morphological evolution through cis-regulatory changes in a pleiotropic gene. Nature 440, 1050–1053 (2006)

    Article  ADS  Google Scholar 

  26. Tao, H., Cox, D. R. & Frazer, K. A. Allele-specific KRT1 expression is a complex trait. PLoS Genet. 2, e93 (2006)

    Article  Google Scholar 

  27. Harr, B., Weiss, S., David, J. R., Brem, G. & Schlötterer, C. A microsatellite-based multilocus phylogeny of the Drosophila melanogaster species complex. Curr. Biol. 8, 1183–1186 (1998)

    Article  CAS  Google Scholar 

  28. Powell, J. R. Progress and Prospects in Evolutionary Biology: The Drosophila Model (Oxford Univ. Press, New York, 1997)

    Google Scholar 

  29. Thummel, C. S. & Pirrotta, V. New pCaSpeR P element vectors. Drosophila Info. Serv. 71, 150 (1992)

    Google Scholar 

  30. Sharma, Y., Cheung, U., Larsen, E. W. & Eberl, D. F. PPTGAL, a convenient Gal4 P-element vector for testing expression of enhancer fragments in Drosophila. Genesis 34, 115–118 (2002)

    Article  CAS  Google Scholar 

  31. Robertson, H. M. et al. A stable genomic source of P element transposase in Drosophila melanogaster. Genetics 118, 461–470 (1988)

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Rubin, G. M. & Spradling, A. C. Genetic transformation of Drosophila with transposable element vectors. Science 218, 348–353 (1982)

    Article  ADS  CAS  Google Scholar 

  33. Roch, F., Alonso, C. R. & Akam, M. Drosophila miniature and dusky encode ZP proteins required for cytoskeletal reorganisation during wing morphogenesis. J. Cell Sci. 116, 1199–1207 (2003)

    Article  CAS  Google Scholar 

  34. Patel, N. H. et al. Expression of engrailed proteins in arthropods, annelids, and chordates. Cell 58, 955–968 (1989)

    Article  CAS  Google Scholar 

  35. Blochlinger, K., Bodmer, R., Jan, L. Y. & Jan, Y. N. Patterns of expression of cut, a protein required for external sensory organ development in wild-type and cut mutant Drosophila embryos. Genes Dev. 4, 1322–1331 (1990)

    Article  CAS  Google Scholar 

  36. True, J. R., Mercer, J. M. & Laurie, C. C. Differences in crossover frequency and distribution among three sibling species of Drosophila. Genetics 142, 507–523 (1996)

    CAS  PubMed  PubMed Central  Google Scholar 

  37. Gleason, J. M., Cropp, K. A. & Dewoody, R. S. DNA preparations from fly wings for molecular marker assisted crosses. Drosophila Info. Serv. 87, 107–108 (2004)

    Google Scholar 

  38. Stern, D. L. & Sucena, E. in Drosophila: A Laboratory Manual (eds Ashburner, M., Hawley, S. & Sullivan, B.) 601–615 (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, 2000)

    Google Scholar 

Download references


We thank Y. Tao for D. mauritiana stocks and for sharing unpublished data; F. Roch for anti-Min antibody; T. Frankino for help with flies; A. Bassan, P. Valenti, Y. Latapie and S. Plaza for experimental help, discussions and critical reading of the manuscript; and W. Damen and N. Brown for hosting A.P.M. and I.D., respectively, during manuscript revisions. This work was supported by funding from the Association pour la Recherche sur le Cancer (to J.Z.), the Fondation pour la Recherche Médicale (programme équipe 2005), an EMBO Long-Term Fellowship and a BBSRC grant to I.D., a Damon Runyon Cancer Research Foundation Fellowship to V.O., a National Institute of General Medical Sciences National Research Service Award Fellowship to D.G.S., and a NIH grant and a David and Lucile Packard Foundation Fellowship to D.L.S.

Author Contributions The enhancer analysis was designed by A.P.M., I.D., F.P. and D.L.S.; DNA constructs and transgenic flies were made by A.P.M., I.D. and D.G.S.; embryos were stained, examined and photographed by A.P.M., I.D., J.Z., D.G.S., F.P. and D.L.S.; the recombination mapping experiment was designed and performed by D.L.S., V.O. and A.P.M. All authors participated in data analysis and writing of the manuscript.

Author information

Authors and Affiliations


Corresponding author

Correspondence to David L. Stern.

Ethics declarations

Competing interests

Reprints and permissions information is available at The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures S1-S6 with legends and Supplementary Tables S1-S3 (PDF 5038 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

McGregor, A., Orgogozo, V., Delon, I. et al. Morphological evolution through multiple cis-regulatory mutations at a single gene. Nature 448, 587–590 (2007).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.


Quick links

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