Abstract
Protamines are the major DNA-binding proteins in the nucleus of sperm in most vertebrates1,2,3,4,5 and package the DNA in a volume less than 5% of a somatic cell nucleus6. Many mammals have one protamine, but a few species, including humans and mice, have two7,8,9,10,11,12,13. Here we use gene targeting to determine if the second protamine provides redundancy to an essential process, or if both protamines are necessary. We disrupted the coding sequence of one allele of either Prm1 or Prm2 in embryonic stem (ES) cells derived from 129-strain mice, and injected them into blastocysts from C57BL/6-strain mice. Male chimeras produced 129-genotype sperm with disrupted Prm1 or Prm2 alleles, but failed to sire offspring carrying the 129 genome. We also found that a decrease in the amount of either protamine disrupts nuclear formation, processing of protamine-2 and normal sperm function. Our studies show that both protamines are essential and that haploinsufficiency caused by a mutation in one allele of Prm1 or Prm2 prevents genetic transmission of both mutant and wild-type alleles.
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References
Hecht, N.B. Mammalian protamines and their expression. in Histones and Other Basic Nuclear Proteins 347–374 (CRC, Orlando, 1989).
Meistrich, M.L. Histone and basic nuclear protein transitions in mammalian spermatogenesis. in Histones and Other Basic Nuclear Proteins 165–182 (CRC, Orlando, 1989).
Balhorn, R. Mammalian protamines: structure and molecular interactions. in Molecular Biology of Chromosome Function 366–395 (Springer, New York, 1989).
Hecht, N.B. Regulation of 'haploid expressed genes' in male germ cells. J. Reprod. Fertil. 88, 679–693 (1990).
Balhorn, R. et al. Protamine mediated condensation of DNA in mammalian sperm. in The Male Gamete: From Basic Science to Clinical Applications 55–70 (Cache River, Viena, Illinois, 1999).
Wyrobek, A.J., Meistrich, M.L., Furrer, R. & Bruce, W.R. Physical characteristics of mouse sperm nuclei. Biophys. J. 16, 811–825 (1976).
Bellvé, A.R., Anderson, E. & Hanley-Bowdoin, L. Synthesis and amino acid composition of basic proteins in mammalian sperm nuclei. Dev. Biol. 47, 349–365 (1975).
Balhorn, R., Gledhill, B.L. & Wyrobek, A.J. Mouse sperm chromatin proteins: quantitative isolation and partial characterization. Biochemistry 16, 4074–4080 (1977).
Kleene, K.C., Distel, R.J. & Hecht, N.B. Nucleotide sequence of a cDNA clone encoding mouse protamine 1. Biochemistry 24, 719–722 (1985).
McKay, D.J., Renaux, B.S. & Dixon, G.H. The amino acid sequence of human sperm protamine P1. Biosci. Rep. 5, 383–391 (1985).
Yelick, P.C. et al. Mouse protamine 2 is synthesized as a precursor whereas mouse protamine 1 is not. Mol. Cell. Biol. 7, 2173–2179 (1987).
Domenjoud, L., Fronia, C., Uhde, F. & Engel, W. Sequence of human protamine 2 cDNA. Nucleic Acids Res. 16, 7733 (1988).
Oliva, R. & Dixon, G.H. Vertebrate protamine genes and the histone-to-protamine replacement reaction. Prog. Nucleic Acids Res. Mol. Biol. 40, 25–94 (1991).
Cho, C., Myles, D.G. & Primakoff, P. A PCR method for distinguishing cells from mouse strains 129 and C57BL/6 for gene knockout studies. Technical Tips Online 1:20:T01139 (1997). http://research.bmn.com/tto/
Kosower, N.S., Katayose, H. & Yanagimachi, R. Thiol-disulfide status and acridine orange fluorescence of mammalian sperm nuclei. J. Androl. 13, 342–348 (1992).
Yu, Y.E. et al. Abnormal spermatogenesis and reduced fertility in transition nuclear protein 1-deficient mice. Proc. Natl. Acad. Sci. USA 97, 4683–4688 (2000).
Debarle, M., Martinage, A., Sautiere, P. & Chevaillier, P. Persistence of protamine precursors in mature sperm nuclei of the mouse. Mol. Reprod. Dev. 40, 84–90 (1995).
Balhorn, R., Weston, S., Thomas, C. & Wyrobek, A.J. DNA packaging in mouse spermatids. Synthesis of protamine variants and four transition proteins. Exp. Cell Res. 150, 298–308 (1984).
Lee, K., Haugen, H.S., Clegg, C.H. & Braun, R.E. Premature translation of protamine 1 mRNA causes precocious nuclear condensation and arrests spermatid differentiation in mice. Proc. Natl. Acad. Sci. USA 92, 12451–12455 (1995).
Wu, J.Y. et al. Spermiogenesis and exchange of basic nuclear proteins are impaired in male germ cells lacking Camk4. Nature Genet. 25, 448–452 (2000).
Kuratani, S., Matsuo, I. & Aizawa, S. Developmental patterning and evolution of the mammalian viscerocranium: genetic insights into comparative morphology. Dev. Dyn. 209, 139–155 (1997).
Fero, M.L., Randel, E., Gurley, K.E., Roberts, J.M. & Kemp, C.J. The murine gene p27Kip1 is haplo-insufficient for tumor suppression. Nature 396, 177–180 (1998).
Braun, R.E. et al. Genetically haploid spermatids are phenotypically diploid. Nature 337, 373–376 (1989).
Caldwell, K.A. & Handel, M.A. Protamine transcript sharing among postmeiotic spermatids. Proc. Natl. Acad. Sci. USA 88, 2407–2411 (1991).
Mbikay, M. et al. Impaired fertility in mice deficient for the testicular germ-cell protease PC4. Proc. Natl. Acad. Sci. USA 94, 6842–6846 (1997).
Balhorn, R., Reed, S. & Tanphaichitr, N. Aberrant protamine 1/protamine 2 ratios in sperm of infertile human males. Experentia 44, 52–55 (1988).
de Yebra, l.L., Ballesca, J.L., Vanrell, J.A., Bassas, L. & Oliva, R. Complete selective absence of protamine-P2 in humans. J. Biol. Chem. 268, 10553–10557 (1993).
Stanker, L.H., Wyrobek, A., Mckeown, C. & Balhorn, R. Identification of the binding site of two monoclonal antibodies to human protamine. Mol. Immunol. 30, 1633–1638 (1993).
Acknowledgements
We thank R. Balhorn for providing antibodies and C. Rogers for help with experiments. Animal care was in accordance with U.S. Public Health Service guidelines for use of animals, and all procedures were approved in advance by the Institutional Animal Care and Use Committee of the National Institute of Environmental Health Sciences.
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Cho, C., Willis, W., Goulding, E. et al. Haploinsufficiency of protamine-1 or -2 causes infertility in mice. Nat Genet 28, 82–86 (2001). https://doi.org/10.1038/ng0501-82
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DOI: https://doi.org/10.1038/ng0501-82
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