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.

Genomic and functional evolution of the Drosophila melanogaster sperm proteome


In addition to delivering a haploid genome to the egg, sperm have additional critical functions, including egg activation, origination of the zygote centrosome and delivery of paternal factors1,2. Despite this, existing knowledge of the molecular basis of sperm form and function is limited. We used whole-sperm mass spectrometry to identify 381 proteins of the Drosophila melanogaster sperm proteome (DmSP). This approach identified mitochondrial, metabolic and cytoskeletal proteins, in addition to several new functional categories. We also observed nonrandom genomic clustering of sperm genes and underrepresentation on the X chromosome. Identification of widespread functional constraint on the proteome indicates that sexual selection has had a limited role in the overall evolution of D. melanogaster sperm. The relevance of the DmSP to the study of mammalian sperm function and fertilization mechanisms is demonstrated by the identification of substantial homology between the DmSP and proteins of the mouse axoneme accessory structure.

This is a preview of subscription content

Access options

Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Drosophila melanogaster sperm purification and 2D gel electrophoresis.
Figure 2: Molecular functional composition of the DmSP based on Gene Ontology annotation.
Figure 3: Genome distribution of the DmSP.
Figure 4: Evolutionary analysis of the DmSP.


  1. Loppin, B. & Karr, T.L. Molecular genetics of insect fertilization. in Comprehensive Insect Molecular Science (eds. Gilbert, L.B. & Iatrou, K.) (Elsevier, Oxford, 2004).

    Google Scholar 

  2. Loppin, B., Lepetit, D., Dorus, S., Couble, P. & Karr, T.L. Origin and neofunctionalization of a Drosophila paternal effect gene essential for zygote viability. Curr. Biol. 15, 87–93 (2005).

    CAS  Article  Google Scholar 

  3. Snook, R.R., Cleland, S.Y., Wolfner, M.F. & Karr, T.L. Offsetting effects of Wolbachia infection and heat shock on sperm production in Drosophila simulans: analyses of fecundity, fertility and accessory gland proteins. Genetics 155, 167–178 (2000).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Mitchell, B.F., Pedersen, L.B., Feely, M., Rosenbaum, J.L. & Mitchell, D.R. ATP production in Chlamydomonas reinhardtii flagella by glycolytic enzymes. Mol. Biol. Cell 16, 4509–4518 (2005).

    CAS  Article  Google Scholar 

  5. Fernandez, D., Valdivia, A., Irazusta, J., Ochoa, C. & Casis, L. Peptidase activities in human semen. Peptides 23, 461–468 (2002).

    CAS  Article  Google Scholar 

  6. Togo, T. & Morisawa, M. GPI-anchored aminopeptidase is involved in the acrosome reaction in sperm of the mussel mytilusedulis. Mol. Reprod. Dev. 67, 465–471 (2004).

    CAS  Article  Google Scholar 

  7. Parsch, J., Meiklejohn, C.D., Hauschteck-Jungen, E., Hunziker, P. & Hartl, D.L. Molecular evolution of the ocnus and janus genes in the Drosophila melanogaster species subgroup. Mol. Biol. Evol. 18, 801–811 (2001).

    CAS  Article  Google Scholar 

  8. Yeh, S.D. et al. Isolation and properties of Gas8, a growth arrest-specific gene regulated during male gametogenesis to produce a protein associated with the sperm motility apparatus. J. Biol. Chem. 277, 6311–6317 (2002).

    CAS  Article  Google Scholar 

  9. Challapalli, K.K. et al. High reproducibility of large-gel two-dimensional electrophoresis. Electrophoresis 25, 3040–3047 (2004).

    CAS  Article  Google Scholar 

  10. Perotti, M.E., Cattaneo, F., Pasini, M.E., Verni, F. & Hackstein, J.H. Male sterile mutant casanova gives clues to mechanisms of sperm-egg interactions in Drosophila melanogaster. Mol. Reprod. Dev. 60, 248–259 (2001).

    CAS  Article  Google Scholar 

  11. Nurminsky, D.I., Nurminskaya, M.V., De Aguiar, D. & Hartl, D.L. Selective sweep of a newly evolved sperm-specific gene in Drosophila. Nature 396, 572–575 (1998).

    CAS  Article  Google Scholar 

  12. Lu, A.Q. & Beckingham, K. Androcam, a Drosophila calmodulin-related protein, is expressed specifically in the testis and decorates loop kl-3 of the Y chromosome. Mech. Dev. 94, 171–181 (2000).

    CAS  Article  Google Scholar 

  13. Cao, W., Gerton, G.L. & Moss, S.B. Proteomic profiling of accessory structures from the mouse sperm flagellum. Mol. Cell. Proteomics 5, 801–810 (2006).

    CAS  Article  Google Scholar 

  14. Jamieson, B.G.M., Dallai, R. & Afzelius, B.A. Insects: Their Spermatozoa and Phylogeny (Science Publishers, Enfield, New Hampshire, 1999).

    Google Scholar 

  15. Parisi, M. et al. A survey of ovary-, testis-, and soma-biased gene expression in Drosophila melanogaster adults. Genome Biol. 5, R40 (2004).

    Article  Google Scholar 

  16. Parisi, M. et al. Paucity of genes on the Drosophila X chromosome showing male-biased expression. Science 299, 697–700 (2003).

    CAS  Article  Google Scholar 

  17. Boutanaev, A.M., Kalmykova, A.I., Shevelyov, Y.Y. & Nurminsky, D.I. Large clusters of co-expressed genes in the Drosophila genome. Nature 420, 666–669 (2002).

    CAS  Article  Google Scholar 

  18. Lercher, M.J., Urrutia, A.O. & Hurst, L.D. Clustering of housekeeping genes provides a unified model of gene order in the human genome. Nat. Genet. 31, 180–183 (2002).

    CAS  Article  Google Scholar 

  19. Swanson, W.J. & Vacquier, V.D. The rapid evolution of reproductive proteins. Nat. Rev. Genet. 3, 137–144 (2002).

    CAS  Article  Google Scholar 

  20. Fay, J.C., Wyckoff, G.J. & Wu, C.I. Testing the neutral theory of molecular evolution with genomic data from Drosophila. Nature 415, 1024–1026 (2002).

    CAS  Article  Google Scholar 

  21. Swanson, W.J., Clark, A.G., Waldrip-Dail, H.M., Wolfner, M.F. & Aquadro, C.F. Evolutionary EST analysis identifies rapidly evolving male reproductive proteins in Drosophila. Proc. Natl. Acad. Sci. USA 98, 7375–7379 (2001).

    CAS  Article  Google Scholar 

  22. Mueller, J.L. et al. Cross-species comparison of Drosophila male accessory gland protein genes. Genetics 171, 131–143 (2005).

    CAS  Article  Google Scholar 

  23. Duret, L. & Mouchiroud, D. Determinants of substitution rates in mammalian genes: expression pattern affects selection intensity but not mutation rate. Mol. Biol. Evol. 17, 68–74 (2000).

    CAS  Article  Google Scholar 

  24. Clark, N.L. & Swanson, W.J. Pervasive adaptive evolution in primate seminal proteins. PLoS Genet. 1, 335–342 (2006).

    Google Scholar 

  25. Ficarro, S.B. et al. Phosphoproteome analysis by mass spectrometry and its application to Saccharomyces cerevisiae. Nat. Biotechnol. 20, 301–305 (2002).

    CAS  Article  Google Scholar 

  26. Yates, J.R., III, Eng, J.K., McCormack, A.L. & Schieltz, D. Method to correlate tandem mass spectra of modified peptides to amino acid sequences in the protein database. Anal. Chem. 67, 1426–1436 (1995).

    CAS  Article  Google Scholar 

  27. Pappin, D.J., Hojrup, P. & Bleasby, A.J. Rapid identification of proteins by peptide-mass fingerprinting. Curr. Biol. 3, 327–332 (1993).

    CAS  Article  Google Scholar 

  28. Zheng, Y., Jung, M.K. & Oakley, B.R. Gamma-tubulin is present in Drosophila melanogaster and Homo sapiens and is associated with the centrosome. Cell 65, 817–823 (1991).

    CAS  Article  Google Scholar 

  29. Sardiello, M., Licciulli, F., Catalano, D., Attimonelli, M. & Caggese, C. MitoDrome: a database of Drosophila melanogaster nuclear genes encoding proteins targeted to the mitochondrion. Nucleic Acids Res. 31, 322–324 (2003).

    CAS  Article  Google Scholar 

  30. Li, W.H. Unbiased estimation of the rates of synonymous and nonsynonymous substitution. J. Mol. Evol. 36, 96–99 (1993).

    CAS  Article  Google Scholar 

Download references


We thank S. Pitnick for assistance with sperm isolation and purification and C. Bergman, L. Hurst, J. Fay and B. Heath for discussions and constructive comments concerning the manuscript. We would also like to thank Z.N. Freeman, M. Karr, J. MacNaughton and D. Knowles for technical support. We also acknowledge the expertise of R. Reed at the University of California San Francisco protein sequencing facility. This work was funded in part by a Wolfson-Royal Society Merit Award, the Biotechnology and Biosciences Research Council (BBC0076701) and the US National Science Foundation (MCB-0135166) to T.L.K. and a Ruth L. Kirschstein National Research Service Award to S.D.

Author information

Authors and Affiliations



This study was conceived and initiated by T.L.K. at the University of Chicago; protein identification was performed by S.A.B., J.S., D.F.H and U.G.; bioinformatic and evolutionary analyses were performed by S.D. and T.L.K.; S.D. and T.L.K. contributed to the writing of the paper.

Corresponding author

Correspondence to Timothy L Karr.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

ESI-MS MS spectra of a single hit assignment and deduced amino acid sequence for D. melanogaster gene CG31644. (PDF 208 kb)

Supplementary Table 1

Genes encoding the Drosophila melanogaster sperm proteome. (PDF 268 kb)

Supplementary Table 2

Peptides identified by mass spectrometry. (PDF 361 kb)

Supplementary Table 3

Membrane, signaling and neural genes of the DmSP. (PDF 78 kb)

Supplementary Table 4

2D gel/MALDI-TOF identification of D. melanogaster sperm proteome. (PDF 61 kb)

Supplementary Table 5

Homologous mouse sperm flagellum accessory structure proteins. (PDF 67 kb)

Supplementary Table 6

DmSP evolutionary rates. (PDF 75 kb)

Supplementary Table 7

10% of DmSP genes with the highest Ka/Ks ratios. (PDF 25 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Dorus, S., Busby, S., Gerike, U. et al. Genomic and functional evolution of the Drosophila melanogaster sperm proteome. Nat Genet 38, 1440–1445 (2006).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

Further reading


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