Animal Reproduction (AR)
Animal Reproduction (AR)

What is known so far about bull sperm protamination: a review

Carlos Alonso Paco Nagaki; Thais Rose dos Santos Hamilton; Mayra Elena Ortiz D Ávila Assumpção

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Sperm routinary fitness evaluation is not sufficient to predict bull reproductive capacity as they present differences in fertility up to 40%. Among the defects which compromise spermatozoa functionality, new approaches consider the study of sperm chromatin, which is the core structure containing paternal genetic information. Sperm chromatin needs to be compacted to maintain the integrity of DNA, which occurs by binding nucleoproteins with high affinity to DNA. In the last stages of sperm maturation, chromatin is hyper-compacted by basic proteins called protamines in a process named protamination. In this review, we summarized intrinsic and extrinsic factors that are suggested to influence protamination in bull spermatozoa, considering old and new evidence from human and murine spermatozoa. Also, the current approaches to evaluate bull protamination and its relationship with fertility were described. Nevertheless, the physiological mechanisms of protamination are still poorly understood.


protamine, spermatozoa, DNA fragmentation, spermatogenesis, bull fertility


Absalan F, Movahedin M, Mowla SJ. Evaluation of apoptotic genes expression and its protein after treatment of cryptorchid mice. Iran Biomed J. 2012;16(2):77-83. PMid:22801280.

Agarwal A, Saleh RA, Bedaiwy MA. Role of reactive oxygen species in the pathophysiology of human reproduction. Fertil Steril. 2003;79(4):829-43. PMid:12749418.

Alahmar A. Role of oxidative stress in male infertility: an updated review. J Hum Reprod Sci. 2019;12(1):4-18. PMid:31007461.

Amor H, Zeyad A, Bakry MS, Bosilah AM, Ali HB, Hammadeh ME. Protamine ratio as predictor of the fertility potential of sperm by couple undergoing ICSI. Int J Women’s Health Reprod Sci. 2018;6(4):400-9.

Aoki VW, Emery BR, Liu L, Carrell DT. Protamine levels vary between individual sperm cells of infertile human males and correlate with viability and DNA integrity. J Androl. 2006;27(6):890-8. PMid:16870950.

Aoki VW, Moskovtsev SI, Willis J, Liu L, Mullen JBM, Carrell DT. DNA integrity is compromised in protamine-deficient human sperm. J Androl. 2005;26(6):741-8. PMid:16291969.

Arévalo L, Tourmente M, Varea-Sánchez M, Ortiz-García D, Roldan ERS. Sexual selection towards a protamine expression ratio optimum in two rodent groups? Evolution. 2021;75(8):2124-31. PMid:34224143.

Ausio J, Zhang Y, Ishibashi T. Histone variants and posttranslational modifications in spermatogenesis and infertility. In: García-Giménez JL, editor. Epigenetic biomarkers and diagnostics. London: Elsevier; 2016.

Balhorn R, Brewer L, Corzett M. DNA condensation by protamine and arginine-rich peptides: analysis of toroid stability using single DNA molecules. Mol Reprod Dev. 2000;56(2, Suppl):230-4.<230::AID-MRD3>3.0.CO;2-V. PMid:10824973.

Balhorn R, Reed S, Tanphaichitr N. Aberrant protamine 1/protamine 2 ratios in sperm of infertile human males. Experientia. 1988;44(1):52-5. PMid:3350120.

Balhorn R. The protamine family of sperm nuclear proteins. Genome Biol. 2007;8(9):227. PMid:17903313.

Bao J, Bedford MT. Epigenetic regulation of the histone-to-protamine transition during spermiogenesis. Reproduction. 2016;151(5):R55-70. PMid:26850883.

Belhadj Slimen I, Najar T, Ghram A, Abdrrabba M. Heat stress effects on livestock: Molecular, cellular and metabolic aspects, a review. J Anim Physiol Anim Nutr (Berl). 2016;100(3):401-12. PMid:26250521.

Bennetts LE, Aitken RJ. A comparative study of oxidative DNA damage in mammalian spermatozoa. Mol Reprod Dev. 2005;71(1):77-87. PMid:15736137.

Bissonnette N, Lévesque-Sergerie JP, Thibault C, Boissonneault G. Spermatozoal transcriptome profiling for bull sperm motility: A potential tool to evaluate semen quality. Reproduction. 2009;138(1):65-80. PMid:19423662.

Björndahl L, Kvist U. Human sperm chromatin stabilization: a proposed model including zinc bridges. Mol Hum Reprod. 2010;16(1):23-9. PMid:19933313.

Blendy JA, Kaestner KH, Weinbauer GF, Nieschlag E, Schütz G. Severe impairment of spermatogenesis in mice lacking the CREM gene. Nature. 1996;380(6570):162-5. PMid:8600391.

Bollwein H, Bittner L. Impacts of oxidative stress on bovine sperm function and subsequent in vitro embryo development. Anim Reprod. 2018;15(Suppl 1):703-10. PMid:36249836.

Carreira JT, Trevizan JT, Kipper BH, Perri SHV, Carvalho IR, Rodrigues LH, Silva C, Koivisto MB. Impaired protamination and sperm DNA damage in a Nellore bull with high percentages of morphological sperm defects in comparison to normospermic bulls. Arq Bras Med Vet Zootec. 2015;67(2):417-23.

Castro LS, Siqueira AFP, Hamilton TRS, Mendes CM, Visintin JA, Assumpção MEOA. Effect of bovine sperm chromatin integrity evaluated using three different methods on in vitro fertility. Theriogenology. 2018;107:142-8. PMid:29154161.

Cho C, Jung-Ha H, Willis WD, Goulding EH, Stein P, Xu Z, Schultz RM, Hecht NB, Eddy EM. Protamine 2 deficiency leads to sperm DNA damage and embryo death in mice. Biol Reprod. 2003;69(1):211-7. PMid:12620939.

Choi HY, Kim SK, Kim SH, Choi YM, Jee BC. Impact of sperm DNA fragmentation on clinical in vitro fertilization outcomes. Clin Exp Reprod Med. 2017;44(4):224-31. PMid:29376020.

Codrington AM, Hales BF, Robaire B. Exposure of male rats to cyclophosphamide alters the chromatin structure and basic proteome in spermatozoa. Hum Reprod. 2007;22(5):1431-42. PMid:17303633.

Dadoune J. Expression of mammalian spermatozoal nucleoproteins. Microsc Res Tech. 2003;61(1):56-75. PMid:12672123.

Daly J, Smith H, McGrice HA, Kind KL, van Wettere WHEJ. Towards improving the outcomes of assisted reproductive technologies of cattle and sheep, with particular focus on recipient management. Animals. 2020;10(2):1-15. PMid:32069818.

DeJarnette JM. The effect of semen quality on reproductive efficiency. Vet Clin North Am Food Anim Pract. 2005;21(2):409-18. PMid:15955437.

Dogan S, Vargovic P, Oliveira R, Belser LE, Kaya A, Moura A, Sutovsky P, Parrish J, Topper E, Memili E. Sperm Protamine-Status Correlates to the Fertility of Breeding Bulls1. Biol Reprod. 2015;92(4):92. PMid:25673563.

Eid LN, Lorton SF, Parrish JJ. Paternal influence on S-phase in the first cell cycle of the bovine embryo. Biol Reprod. 1994;51(6):1232-7. PMid:7888500.

Enciso M, Cisale H, Johnston SD, Sarasa J, Fernández JL, Gosálvez J. Major morphological sperm abnormalities in the bull are related to sperm DNA damage. Theriogenology. 2011;76(1):23-32. PMid:21529921.

England K, O’Driscoll C, Cotter TG. Carbonylation of glycolytic proteins is a key response to drug-induced oxidative stress and apoptosis. Cell Death Differ. 2004;11(3):252-60. PMid:14631408.

Erkek S, Hisano M, Liang CY, Gill M, Murr R, Dieker J, Schübeler D, van der Vlag J, Stadler MB, Peters AH. Molecular determinants of nucleosome retention at CpG-rich sequences in mouse spermatozoa. Nat Struct Mol Biol. 2013;20(7):868-75. PMid:23770822.

Evenson DP, Wixon R. Clinical aspects of sperm DNA fragmentation detection and male infertility. Theriogenology. 2006;65(5):979-91. PMid:16242181.

Fatehi AN, Bevers MM, Schoevers E, Roelen BAJ, Colenbrander B, Gadella BM. DNA damage in bovine sperm does not block fertilization and early embryonic development but induces apoptosis after the first cleavages. J Androl. 2006;27(2):176-88. PMid:16304212.

Feugang JM, Rodriguez-Osorio N, Kaya A, Wang H, Page G, Ostermeier GC, Topper EK, Memili E. Transcriptome analysis of bull spermatozoa: implications for male fertility. Reprod Biomed Online. 2010;21(3):312-24. PMid:20638337.

Flowers WL. Triennial Reproduction Symposium: sperm characteristics that limit success of fertilization. J Anim Sci. 2013;91(7):3022-9. PMid:23307855.

Fortes MRS, Satake N, Corbet DH, Corbet NJ, Burns BM, Moore SS, Boe-Hansen GB. Sperm protamine deficiency correlates with sperm DNA damage in Bos indicus bulls. Andrology. 2014;2(3):370-8. PMid:24634207.

Fraga CG, Motchnik PA, Wyrobek AJ, Rempel DM, Ames BN. Smoking and low antioxidant levels increase oxidative damage to sperm DNA. Mutat Res. 1996;351(2):199-203. PMid:8622715.

Freundl G, Grimm HJ, Hofmann N. Selective filtration of abnormal spermatozoa by the cervical mucus*. Hum Reprod. 1988;3(3):277-80. PMid:3372692.

Gao Y, Jian L, Lu W, Xue Y, Machaty Z, Luo H. Vitamin E can promote spermatogenesis by regulating the expression of proteins associated with the plasma membranes and protamine biosynthesis. Gene. 2021;773:145364. PMid:33359122.

Garcia-Oliveros LN, Arruda RP, Batissaco L, Gonzaga VHG, Nogueira VJM, Florez-Rodriguez SA, Almeida FS, Alves MBR, Pinto SCC, Nichi M, Losano JDA, Kawai GKV, Celeghini ECC. Heat stress effects on bovine sperm cells : a chronological approach to early findings. Int J Biometeorol. 2020;64(8):1367-78. PMid:32388687.

Gosálvez Berenguer J, Caballero Peregrín P, López-Fernández C, Fernández JL, Núñez Calonge R. Fragmentación del ADN espermático. Rev Int Androl. 2008;6(3):193-209.

Greco E, Iacobelli M, Rienzi L, Ubaldi F, Ferrero S, Tesarik JAN. Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment. J Androl. 2005;26(3):349-53. PMid:15867002.

Hamidian S, Talebi AR, Fesahat F, Bayat M, Mirjalili AM, Ashrafzadeh HR, Rajabi M, Montazeri F, Babaei S. The effect of vitamin C on the gene expression profile of sperm protamines in the male partners of couples with recurrent pregnancy loss: a randomized clinical trial. Clin Exp Reprod Med. 2020;47(1):68-76. PMid:32146776.

Hamilton TRDS, Siqueira AFP, Castro LS, Mendes CM, Delgado JDC, Assis PM, Mesquita LP, Maiorka PC, Nichi M, Goissis MD, Visintin JA, Assumpção MEODA. Effect of heat stress on sperm DNA: protamine assessment in ram spermatozoa and testicle. Oxid Med Cell Longev. 2018;2018:5413056. PMid:29765499.

Hamilton TRS, Simões R, Assumpção MEODA. An improved acetic acid-urea polyacrylamide electrophoresis method to evaluate bovine sperm protamines. Reprod Domest Anim. 2021;56(7):1050-6. PMid:33890330.

Hamilton TRS, Simões R, Mendes CM, Goissis MD, Nakajima E, Martins EAL, Visintin JA, Assumpção MEOA. Detection of protamine 2 in bovine spermatozoa and testicles. Andrology. 2019;7(3):373. PMid:30920782.

Hammoud S, Emery BR, Aoki VW, Carrell DT. Identification of genetic variation in the 5′ and 3′ non-coding regions of the protamine genes in patients with protamine deregulation. Arch Androl. 2007;53(5):267-74. PMid:18309899.

Hammoud SS, Nix DA, Zhang H, Purwar J, Carrell DT, Cairns BR. Distinctive chromatin in human sperm packages genes for embryo development. Nature. 2009;460(7254):473-8. PMid:19525931.

Harayama H, Minami K, Kishida K, Noda T. Protein biomarkers for male artificial insemination subfertility in bovine spermatozoa. Reprod Med Biol. 2017;16(2):89-98. PMid:29259456.

Hikim APS, Lue Y, Yamamoto CM, Vera Y, Rodriguez S, Yen PH, Soeng K, Wang C, Swerdloff RS. Key apoptotic pathways for heat-induced programmed germ cell death in the testis. Endocrinology. 2003;144(7):3167-75. PMid:12810573.

Hlady RA, Robertson KD. Use of chromatin changes as biomarkers. In: Binda O, Fernandez-Zapico ME, editors. Chromatin signaling and diseases. London: Elsevier; 2016.

Inchaisri C, Jorritsma R, Vos PLAM, van der Weijden GC, Hogeveen H. Economic consequences of reproductive performance in dairy cattle. Theriogenology. 2010;74(5):835-46. PMid:20580069.

Jung YH, Sauria MEG, Lyu X, Cheema MS, Ausio J, Taylor J, Corces VG. Chromatin states in mouse sperm correlate with embryonic and adult regulatory landscapes. Cell Rep. 2017;18(6):1366-82. PMid:28178516.

Kastelic JP. Male involvement in fertility and factors affecting semen quality in bulls. Anim Front. 2013;3(4):20-5.

Kidder HE, Black WG, Wiltbank JN, Ulberg LC, Casida LE. Fertilization Rates and Embryonic Death Rates in Cows Bred to Bulls of Different Levels of Fertility. J Dairy Sci. 1954;37(6):691-7.

Kiefer H, Sellem E, Bonnet-Garnier A, Pannetier M, Costes V, Schibler L, Jammes H. The epigenome of male germ cells and the programming of phenotypes in cattle. Anim Front. 2021;11(6):28-38. PMid:34934527.

Kim B, Park K, Rhee K. Heat stress response of male germ cells. Cell Mol Life Sci. 2013;70(15):2623-36. PMid:23007846.

Kleene KC. Patterns, mechanisms, and functions of translation regulation in mammalian spermatogenic cells. Cytogenet Genome Res. 2003;103(3-4):217-24. PMid:15051942.

Kutchy NA, Velho A, Menezes ESB, Jacobsen M, Thibaudeau G, Wills RW, Moura A, Kaya A, Perkins A, Memili E. Testis specific histone 2B is associated with sperm chromatin dynamics and bull fertility: a pilot study. Reprod Biol Endocrinol. 2017;15(1):59. PMid:28764714.

Larson JL, Miller DJ. Can relative spermatozoal galactosyltransferase activity be predictive of dairy bull fertility? J Dairy Sci. 2000;83(11):2473-9. PMid:11104266.

Lee K, Fajardo MA, Braun RE. A testis cytoplasmic RNA-binding protein that has the properties of a translational repressor. Mol Cell Biol. 1996;16(6):3023-34. PMid:8649414.

Lee K, Haugen HS, Clegg CH, Braun RE. Premature translation of protamine 1 mRNA causes precocious nuclear condensation and arrests spermatid differentiation in mice. Proc Natl Acad Sci USA. 1995;92(26):12451-5. PMid:8618919.

Lewis JD, Abbott DW, Ausió J. A haploid affair: core histone transitions during spermatogenesis. Biochem Cell Biol. 2003;81(3):131-40. PMid:12897846.

Llavanera M, Ribas-Maynou J, Delgado-Bermúdez A, Recuero S, Muiño R, Hidalgo CO, Tamargo C, Bonet S, Mateo-Otero Y, Yeste M. Sperm chromatin condensation as an in vivo fertility biomarker in bulls: a flow cytometry approach. J Anim Sci Biotechnol. 2021;12(1):115. PMid:34749810.

Maier W, Nussbaum G, Domenjoud L, Klemm U, Engel W. The lack of protamine 2 (P2) in boar and bull spermatozoa is due to mutations within the P2 gene. Nucleic Acids Res. 1990;18(5):1249-54. PMid:2320417.

Meistrich ML, Mohapatra B, Shirley CR, Zhao M. Roles of transition nuclear proteins in spermiogenesis. Chromosoma. 2003;111(8):483-8. PMid:12743712.

Ni K, Spiess AN, Schuppe HC, Steger K. The impact of sperm protamine deficiency and sperm DNA damage on human male fertility: a systematic review and meta-analysis. Andrology. 2016;4(5):789-99. PMid:27231200.

Oleszczuk K, Augustinsson L, Bayat N, Giwercman A, Bungum M. Prevalence of high DNA fragmentation index in male partners of unexplained infertile couples. Andrology. 2013;1(3):357-60. PMid:23596042.

Oliva R, De Mateo S, Estanyol JM. Sperm cell proteomics. Proteomics. 2009;9(4):1004-17. PMid:19212950.

Oliva R. Protamines and male infertility. Hum Reprod Update. 2006;12(4):417-35. PMid:16581810.

Pardede BP, Agil M, Supriatna I. Protamine and other proteins in sperm and seminal plasma as molecular markers of bull fertility. Vet World. 2020;13(3):556-62. PMid:32367964.

Paul C, Murray AA, Spears N, Saunders PTK. A single, mild, transient scrotal heat stress causes DNA damage, subfertility and impairs formation of blastocysts in mice. Reproduction. 2008;136(1):73-84. PMid:18390691.

Pérez-Crespo M, Pintado B, Gutiérrez-Adán A. Scrotal heat stress effects on sperm viability, sperm DNA integrity, and the offspring sex ratio in mice. Mol Reprod Dev. 2008;75(1):40-7. PMid:17474098.

Pugacheva EM, Rivero-Hinojosa S, Espinoza CA, Méndez-Catalá CF, Kang S, Suzuki T, Kosaka-Suzuki N, Robinson S, Nagarajan V, Ye Z, Boukaba A, Rasko JE, Strunnikov AV, Loukinov D, Ren B, Lobanenkov VV. Comparative analyses of CTCF and BORIS occupancies uncover two distinct classes of CTCF binding genomic regions. Genome Biol. 2015;16(1):161. PMid:26268681.

Rahman MB, Kamal MM, Rijsselaere T, Vandaele L, Shamsuddin M, Van Soom A. Altered chromatin condensation of heat-stressed spermatozoa perturbs the dynamics of DNA methylation reprogramming in the paternal genome after in vitro fertilisation in cattle. Reprod Fertil Dev. 2014;26(8):1107-16. PMid:24041366.

Rahman MB, Vandaele L, Rijsselaere T, Maes D, Hoogewijs M, Frijters A, Noordman J, Granados A, Dernelle E, Shamsuddin M, Parrish JJ, Van Soom A. Scrotal insulation and its relationship to abnormal morphology, chromatin protamination and nuclear shape of spermatozoa in Holstein-Friesian and Belgian Blue bulls. Theriogenology. 2011;76(7):1246-57. PMid:21777969.

Rathke C, Baarends WM, Awe S, Renkawitz-Pohl R. Chromatin dynamics during spermiogenesis. Biochim Biophys Acta. 2014;1839(3):155-68. PMid:24091090.

Ravel C, Chantot-Bastaraud S, El Houate B, Berthaut I, Verstraete L, De Larouziere V, Lourenço D, Dumaine A, Antoine JM, Mandelbaum J, Siffroi JP, McElreavey K. Mutations in the protamine 1 gene associated with male infertility. Mol Hum Reprod. 2007;13(7):461-4. PMid:17494104.

Saacke RG. Sperm morphology: its relevance to compensable and uncompensable traits in semen. Theriogenology. 2008;70(3):473-8.

Saito H, Hara K, Kitajima S, Tanemura K. Effect of vitamin E deficiency on spermatogenesis in mice and its similarity to aging. Reprod Toxicol. 2020;98:225-32. PMid:33045311.

Samans B, Yang Y, Krebs S, Sarode GV, Blum H, Reichenbach M, Wolf E, Steger K, Dansranjavin T, Schagdarsurengin U. Uniformity of nucleosome preservation pattern in mammalian sperm and Its connection to repetitive DNA elements. Dev Cell. 2014;30(1):23-35. PMid:24998597.

Schneider S, Balbach M, Jan F Jikeli, Fietz D, Nettersheim D, Jostes S, Schmidt R, Kressin M, Bergmann M, Wachten D, Steger K, Schorle H. Re-visiting the Protamine-2 locus: Deletion, but not haploinsufficiency, renders male mice infertile. Sci Rep. 2016;6(1):36764. PMid:27833122.

Seegers H, Fourichon C, Malher X, L’Hostis M. A framework for animal health management. Vet Res. 1994;25(2-3):165-73. PMid:8038779.

Shamsuddin M, Larsson B. In vitro development of bovine embryos after fertilization using semen from different donors. Reprod Domest Anim. 1993;28(2):77-84.

Shinagawa T, Huynh LM, Takagi T, Tsukamoto D, Tomaru C, Kwak HG, Dohmae N, Noguchi J, Ishii S. Disruption of TH2a and TH2b genes causes defects in spermatogenesis. Development. 2015;142(7):1287-92. PMid:25742800.

Simões R, Feitosa WB, Siqueira AFP, Nichi M, Paula-Lopes FF, Marques MG, Peres MA, Barnabe VH, Visintin JA, Assumpção ME. Influence of bovine sperm DNA fragmentation and oxidative stress on early embryo in vitro development outcome. Reproduction. 2013;146(5):433-41. PMid:23940385.

Souza ET, Silva CV, Travençolo BAN, Alves BG, Beletti ME. Sperm chromatin alterations in fertile and subfertile bulls. Reprod Biol. 2018;18(2):177-81. PMid:29705068.

Steger K, Balhorn R. Sperm nuclear protamines: a checkpoint to control sperm chromatin quality. Anat Histol Embryol. 2018;47(4):273-9. PMid:29797354.

Takeda N, Yoshinaga K, Furushima K, Takamune K, Li Z, Abe SI, Aizawa S, Yamamura K. Viable offspring obtained from Prm1-deficient sperm in mice. Sci Rep. 2016;6(1):27409. PMid:27250771.

Talbert PB, Henikoff S. Histone variants ancient wrap artists of the epigenome. Nat Rev Mol Cell Biol. 2010;11(4):264-75. PMid:20197778.

Vilfan ID, Conwell CC, Hud NV. Formation of native-like mammalian sperm cell chromatin with folded bull protamine. J Biol Chem. 2004;279(19):20088-95. PMid:14990583.

Villani P, Eleuteri P, Grollino MG, Rescia M, Altavista P, Spanò M, Pacchierotti F, Cordelli E. Sperm DNA fragmentation induced by DNAse I and hydrogen peroxide: an in vitro comparative study among different mammalian species. Reproduction. 2010;140(3):445-52. PMid:20584992.

Widayati D. Embryo transfer as an assisted reproductive technology in farm animals. World Acad Sci Eng Technol. 2012;6:10-21.

Wu PY, Scarlata E, O’Flaherty C. Long-term adverse effects of oxidative stress on rat epididymis and spermatozoa. Antioxidants. 2020;9(2):170. PMid:32093059.

Xing W, Krishnamurthy H, Sairam MR. Role of folltropin receptor signaling in nuclear protein transitions and chromatin condensation during spermatogenesis. Biochem Biophys Res Commun. 2003;312(3):697-701. PMid:14680821.

Yu J, Hecht NB, Schultz RM. RNA-binding properties and translation repression in vitro by germ cell-specific MSY2 protein. Biol Reprod. 2002;67(4):1093-8. PMid:12297523.

Yuen BTK, Bush KM, Barrilleaux BL, Cotterman R, Knoepfler PS. Histone H3.3 regulates dynamic chromatin states during spermatogenesis. Development. 2014;141(18):3483-94. PMid:25142466.

Zhu B, Walker SK, Oakey H, Setchell BP, Maddocks S. Effect of paternal heat stress on the development in vitro of preimplantation embryos in the mouse. Andrologia. 2004;36(6):384-94. PMid:15541055.

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