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Sperm chromatin proteomics identifies evolutionarily conserved fertility factors

Nature volume 443, pages 101–105 (2006)Cite this article

Abstract

Male infertility is a long-standing enigma of significant medical concern. The integrity of sperm chromatin is a clinical indicator of male fertility and in vitro fertilization potential1: chromosome aneuploidy and DNA decondensation or damage are correlated with reproductive failure. Identifying conserved proteins important for sperm chromatin structure and packaging can reveal universal causes of infertility. Here we combine proteomics, cytology and functional analysis in Caenorhabditis elegans to identify spermatogenic chromatin-associated proteins that are important for fertility. Our strategy employed multiple steps: purification of chromatin from comparable meiotic cell types, namely those undergoing spermatogenesis or oogenesis; proteomic analysis by multidimensional protein identification technology (MudPIT) of factors that co-purify with chromatin; prioritization of sperm proteins based on abundance; and subtraction of common proteins to eliminate general chromatin and meiotic factors. Our approach reduced 1,099 proteins co-purified with spermatogenic chromatin, currently the most extensive catalogue, to 132 proteins for functional analysis. Reduction of gene function through RNA interference coupled with protein localization studies revealed conserved spermatogenesis-specific proteins vital for DNA compaction, chromosome segregation, and fertility. Unexpected roles in spermatogenesis were also detected for factors involved in other processes. Our strategy to find fertility factors conserved from C. elegans to mammals achieved its goal: of mouse gene knockouts corresponding to nematode proteins, 37% (7/19) cause male sterility. Our list therefore provides significant opportunity to identify causes of male infertility and targets for male contraceptives.

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Figure 1: Abundance-correlated subtractive proteomic strategy to identify spermatogenesis-enriched proteins.
Figure 2: Identification of spermatogenic chromatin-associated proteins by proteomic analysis.

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Acknowledgements

We thank S. Strome, K. Bennett, H.-S. Koo, L. Moore, E. Shulze and J. Aris for providing antibodies; A. Villenueve, G. Stanfield, S. Mitani and the C. elegans Genetic Center (CGC) for providing strains; V. Reinke, L. Moore and R. Navarro for sharing unpublished data; D. King for peptide synthesis; A. Chan for help with microscopy; S. Chu for statistical analysis; A. Severson, T. Cline, A. Skop, E. Xu and J. Gladden for comments on the manuscript; and members of the Meyer laboratory for input on this project. This work was funded by the National Institutes of Health grants to D.S.C., J.R.Y. and B.J.M. B.J.M. is an investigator of the Howard Hughes Medical Institute.

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Authors and Affiliations

  1. Department of Biology, San Francisco State University, 1600 Holloway Avenue, California, 94132, San Francisco, USA

    Diana S. Chu & Tammy F. Wu

  2. Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, California, 92037, La Jolla, USA

    Hongbin Liu & John R. Yates III

  3. Agilent Technologies, Inc., 2850 Centerville Road, Delaware, 19808, Wilmington, USA

    Hongbin Liu

  4. Department of Molecular and Cell Biology, University of California, Berkeley, California, 94720, Berkeley, USA

    Paola Nix, Edward J. Ralston & Barbara J. Meyer

  5. Howard Hughes Medical Institute, California, 94720, Berkeley, USA

    Paola Nix, Edward J. Ralston & Barbara J. Meyer

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  1. Diana S. Chu
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Correspondence to Diana S. Chu.

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Supplementary information

Supplementary Notes

This file contains Supplementary Methods that contain additional detailed methods used in the paper, Supplementary references, legends for Supplementary Figs 1–5 and a legend for Supplementary Table 6. (DOC 112 kb)

Supplementary Figures

This file contains Supplementary figures 1–5. (PDF 1188 kb)

Supplementary Table 1

Abundant spermatogenesis-enriched proteins copurified with chromatin identified by comparative proteomic analysis. (PDF 68 kb)

Supplementary Table 2

Low abundance spermatogenic proteins copurified with chromatin identified by comparative proteomic analysis (PDF 102 kb)

Supplementary Table 3

All shared proteins copurified with chromatin identified by comparative proteomic analysis. (PDF 139 kb)

Supplementary Table 4

Oogenic proteins copurified with chromatin identified by comparative proteomic analysis. (PDF 87 kb)

Supplementary Table 5

This table lists the composition of identified proteins from spermatogenic chromatin samples in functional categories. (PDF 34 kb)

Supplementary Table 6

This table summarizes defects for genes found to be important for fertility by RNAi analysis. (PDF 61 kb)

Supplementary Table 7

This table lists abundant spermatogenesis-enriched chromatin proteins with function in C. elegans fertility. The protein localization has not yet been determined in C. elegans (Category III proteins). Mouse and human homologs with a minimum E value of 1e-9 are listed. (PDF 62 kb)

Supplementary Table 8

The table lists abundant C. elegans spermatogenesis-enriched chromatin proteins homologous to mammalian fertility factors. Mouse and human homologs with a minimum E value of 1e-9 are listed. (PDF 62 kb)

Supplementary Table 9

This table lists abundant C. elegans spermatogenesis-enriched chromatin proteins with mammalian homologs not yet linked to fertility. Mouse and human homologs with a minimum E value of 1e-9 are listed. (PDF 80 kb)

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Chu, D., Liu, H., Nix, P. et al. Sperm chromatin proteomics identifies evolutionarily conserved fertility factors. Nature 443, 101–105 (2006). https://doi.org/10.1038/nature05050

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