Home LiteratureArticle Details
PMID: 31824947 Published · epublish English Journal Article

Proteomic Changes in Human Sperm During Sequential in vitro Capacitation and Acrosome Reaction.

Frontiers in cell and developmental biology ·Vol. 7 ·2019-00-00 ·Pages 295

Castillo J, Bogle OA, Jodar M, Torabi F, Delgado-Dueñas D, Estanyol JM, Ballescà JL, Miller D, Oliva R

Abstract

The male gamete is not completely mature after ejaculation and requires further events in the female genital tract to acquire fertilizing ability, including the processes of capacitation and acrosome reaction. In order to shed light on protein changes experienced by the sperm cell in preparation for fertilization, a comprehensive quantitative proteomic profiling based on isotopic peptide labeling and liquid chromatography followed by tandem mass spectrometry was performed on spermatozoa from three donors of proven fertility under three sequential conditions: purification with density gradient centrifugation, incubation with capacitation medium, and induction of acrosome reaction by exposure to the calcium ionophore A23187. After applying strict selection criteria for peptide quantification and for statistical analyses, 36 proteins with significant changes in their relative abundance within sperm protein extracts were detected. Moreover, the presence of peptide residues potentially harboring sites for post-translational modification was revealed, suggesting that protein modification may be an important mechanism in sperm maturation. In this regard, increased levels of proteins mainly involved in motility and signaling, both regulated by protein modifiers, were detected in sperm lysates following incubation with capacitation medium. In contrast, less abundant proteins in acrosome-reacted cell lysates did not contain potentially modifiable residues, suggesting the possibility that all those proteins might be relocated or released during the process. Protein-protein interaction analysis revealed a subset of proteins potentially involved in sperm maturation, including the proteins Erlin-2 (ERLIN2), Gamma-glutamyl hydrolase (GGH) and Transmembrane emp24 domain-containing protein 10 (TMED10). These results contribute to the current knowledge of the molecular basis of human fertilization. It should now be possible to further validate the potential role of the detected altered proteins as modulators of male infertility.

Keywords
TMT labeling acrosome reaction capacitation mass spectrometry proteomics sperm spermatozoa
Authors & Affiliations
9 authors, click to expand affiliations / ORCID
Castillo Judit
Molecular Biology of Reproduction and Development Research Group, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Fundació Clínic per a la Recerca Biomèdica, Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Universitat de Barcelona, Barcelona, Spain.
Bogle Orleigh Adeleccia
Molecular Biology of Reproduction and Development Research Group, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Fundació Clínic per a la Recerca Biomèdica, Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Universitat de Barcelona, Barcelona, Spain.
Jodar Meritxell
Molecular Biology of Reproduction and Development Research Group, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Fundació Clínic per a la Recerca Biomèdica, Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Universitat de Barcelona, Barcelona, Spain.
Torabi Forough
LIGHT Laboratories, Leeds Institute of Cardiovascular and Metabolic Medicine, University of Leeds, Leeds, United Kingdom.
Delgado-Dueñas David
Molecular Biology of Reproduction and Development Research Group, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Fundació Clínic per a la Recerca Biomèdica, Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Universitat de Barcelona, Barcelona, Spain.
Estanyol Josep Maria
Proteomics Unit, Scientific and Technical Services, Universitat de Barcelona, Barcelona, Spain.
Ballescà Josep Lluís
Clinic Institute of Gynaecology, Obstetrics and Neonatology, Hospital Clínic, Barcelona, Spain.
Miller David
LIGHT Laboratories, Leeds Institute of Cardiovascular and Metabolic Medicine, University of Leeds, Leeds, United Kingdom.
Oliva Rafael
Molecular Biology of Reproduction and Development Research Group, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Fundació Clínic per a la Recerca Biomèdica, Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Universitat de Barcelona, Barcelona, Spain. | Biochemistry and Molecular Genetics Service, Hospital Clínic, Barcelona, Spain.
References (90)
90 references, click to expand
  1. Protamines and male infertility.
    Hum Reprod Update. 2006 Jul-Aug;12(4):417-35 PMID: 16581810
  2. AKAP3 degradation in sperm capacitation is regulated by its tyrosine phosphorylation.
    Biochim Biophys Acta. 2015 Sep;1850(9):1912-20 PMID: 26093290
  3. Advances in sperm proteomics: best-practise methodology and clinical potential.
    Expert Rev Proteomics. 2015 Jun;12(3):255-77 PMID: 25921224
  4. Sperm proteomics: road to male fertility and contraception.
    Int J Endocrinol. 2013;2013:360986 PMID: 24363670
  5. Identification of sperm head proteins involved in zona pellucida binding.
    Hum Reprod. 2013 Apr;28(4):852-65 PMID: 23355646
  6. Identification of proteins involved in human sperm motility using high-throughput differential proteomics.
    J Proteome Res. 2014 Dec 5;13(12):5670-84 PMID: 25250979
  7. Acrosome reaction and Ca²⁺ imaging in single human spermatozoa: new regulatory roles of [Ca²⁺]i.
    Biol Reprod. 2014 Sep;91(3):67 PMID: 25100708
  8. Ca2+ oscillations at fertilization in mammals.
    Biochem Soc Trans. 2003 Oct;31(Pt 5):907-11 PMID: 14505446
  9. Membrane vesiculation as a feature of the mammalian acrosome reaction.
    J Cell Biol. 1967 Sep;34(3):C1-5 PMID: 6050956
  10. CFTR/ENaC-dependent regulation of membrane potential during human sperm capacitation is initiated by bicarbonate uptake through NBC.
    J Biol Chem. 2018 Jun 22;293(25):9924-9936 PMID: 29743243
  11. The presence, role and clinical use of spermatozoal RNAs.
    Hum Reprod Update. 2013 Nov-Dec;19(6):604-24 PMID: 23856356
  12. Dissociation between AKAP3 and PKARII promotes AKAP3 degradation in sperm capacitation.
    PLoS One. 2013 Jul 23;8(7):e68873 PMID: 23894359
  13. Outer dense fibers stabilize the axoneme to maintain sperm motility.
    J Cell Mol Med. 2018 Mar;22(3):1755-1768 PMID: 29168316
  14. Sperm function, protein phosphorylation, and metabolism differ in mice lacking successive sperm-specific glycolytic enzymes.
    Biol Reprod. 2017 Oct 1;97(4):586-597 PMID: 29025010
  15. TMED10 Protein Interferes with Transforming Growth Factor (TGF)-β Signaling by Disrupting TGF-β Receptor Complex Formation.
    J Biol Chem. 2017 Mar 10;292(10):4099-4112 PMID: 28115518
  16. CatSper channel, sperm function and male fertility.
    Reprod Biomed Online. 2015 Jan;30(1):28-38 PMID: 25457194
  17. Loss of R2D2 proteins ROPN1 and ROPN1L causes defects in murine sperm motility, phosphorylation, and fibrous sheath integrity.
    Biol Reprod. 2013 Feb 21;88(2):41 PMID: 23303679
  18. The combined human sperm proteome: cellular pathways and implications for basic and clinical science.
    Hum Reprod Update. 2014 Jan-Feb;20(1):40-62 PMID: 24082039
  19. Statistical analysis of proteomics data: A review on feature selection.
    J Proteomics. 2019 Apr 30;198:18-26 PMID: 30529743
  20. Characteristics of testis-specific phosphoglycerate kinase 2 and its association with human sperm quality.
    Hum Reprod. 2016 Feb;31(2):273-9 PMID: 26677959
  21. Regulation of protein phosphorylation during sperm capacitation.
    Biol Reprod. 1998 Jul;59(1):1-6 PMID: 9674985
  22. In-depth proteomic analysis of the human sperm reveals complex protein compositions.
    J Proteomics. 2013 Feb 21;79:114-22 PMID: 23268119
  23. Identification of several proteins involved in regulation of sperm motility by proteomic analysis.
    Fertil Steril. 2007 Feb;87(2):436-8 PMID: 17074334
  24. Kinases, phosphatases and proteases during sperm capacitation.
    Cell Tissue Res. 2012 Sep;349(3):765-82 PMID: 22427115
  25. Redox regulation of mammalian sperm capacitation.
    Asian J Androl. 2015 Jul-Aug;17(4):583-90 PMID: 25926608
  26. Essential Role of CFTR in PKA-Dependent Phosphorylation, Alkalinization, and Hyperpolarization During Human Sperm Capacitation.
    J Cell Physiol. 2017 Jun;232(6):1404-1414 PMID: 27714810
  27. Isolation and proteomic analysis of mouse sperm detergent-resistant membrane fractions: evidence for dissociation of lipid rafts during capacitation.
    Biol Reprod. 2005 Oct;73(4):721-9 PMID: 15917346
  28. Molecular Basis of Human Sperm Capacitation.
    Front Cell Dev Biol. 2018 Jul 27;6:72 PMID: 30105226
  29. Zona pellucida-binding protein 2 (ZPBP2) and several proteins containing BX7B motifs in human sperm may have hyaluronic acid binding or recognition properties.
    Mol Hum Reprod. 2017 Dec 1;23(12):803-816 PMID: 29126140
  30. Radial spoke protein 44 (human meichroacidin) is an axonemal alloantigen of sperm and cilia.
    Gene. 2007 Jul 1;396(1):93-107 PMID: 17451891
  31. Sedimentation properties in density gradients correspond with levels of sperm DNA fragmentation, chromatin compaction and binding affinity to hyaluronic acid.
    Reprod Biomed Online. 2017 Mar;34(3):298-311 PMID: 28017538
  32. Regulation of Sperm Capacitation and the Acrosome Reaction by PIP 2 and Actin Modulation.
    Asian J Androl. 2015 Jul-Aug;17(4):597-600 PMID: 25966627
  33. The pattern of tyrosine phosphorylation in human sperm in response to binding to zona pellucida or hyaluronic acid.
    Reprod Sci. 2014 May;21(5):573-81 PMID: 24077441
  34. SPFH1 and SPFH2 mediate the ubiquitination and degradation of inositol 1,4,5-trisphosphate receptors in muscarinic receptor-expressing HeLa cells.
    Biochim Biophys Acta. 2009 Nov;1793(11):1710-8 PMID: 19751772
  35. The erlin2 T65I mutation inhibits erlin1/2 complex-mediated inositol 1,4,5-trisphosphate receptor ubiquitination and phosphatidylinositol 3-phosphate binding.
    J Biol Chem. 2018 Oct 5;293(40):15706-15714 PMID: 30135210
  36. The acrosome reaction in human spermatozoa.
    Biol Cell. 2000 Jul;92(3-4):255-66 PMID: 11043413
  37. Comparative proteomic study between human normal motility sperm and idiopathic asthenozoospermia.
    World J Urol. 2013 Dec;31(6):1395-401 PMID: 23455884
  38. Associations between single nucleotide polymorphisms in folate uptake and metabolizing genes with blood folate, homocysteine, and DNA uracil concentrations.
    Am J Clin Nutr. 2008 Oct;88(4):1149-58 PMID: 18842806
  39. Use of phosphoproteomics to study tyrosine kinase activity in capacitating boar sperm. Kinase activity and capacitation.
    Theriogenology. 2005 Jan 15;63(2):599-614 PMID: 15626419
  40. Protein phosphatases decrease their activity during capacitation: a new requirement for this event.
    PLoS One. 2013 Dec 02;8(12):e81286 PMID: 24312544
  41. Characterization of sperm surface protein patterns of ejaculated and capacitated boar sperm, with the detection of ZP binding candidates.
    Int J Biol Macromol. 2013 Oct;61:322-8 PMID: 23916641
  42. Differences between high- and low-motility buffalo sperm identified by comparative proteomics.
    Reprod Domest Anim. 2015 Jun;50(3):443-51 PMID: 25809445
  43. Identification of sperm-specific proteins that interact with A-kinase anchoring proteins in a manner similar to the type II regulatory subunit of PKA.
    J Biol Chem. 2001 May 18;276(20):17332-8 PMID: 11278869
  44. A simple method for assessment of the human acrosome reaction of spermatozoa bound to the zona pellucida: lack of relationship with ionophore A23187-induced acrosome reaction.
    Hum Reprod. 1996 Mar;11(3):551-7 PMID: 8671264
  45. Targeted NGS gene panel identifies mutations in RSPH1 causing primary ciliary dyskinesia and a common mechanism for ciliary central pair agenesis due to radial spoke defects.
    Hum Mol Genet. 2014 Jul 1;23(13):3362-74 PMID: 24518672
  46. Marked differences in protamine content and P1/P2 ratios in sperm cells from percoll fractions between patients and controls.
    J Androl. 2003 May-Jun;24(3):438-47 PMID: 12721220
  47. Identification of protein changes in human spermatozoa throughout the cryopreservation process.
    Andrology. 2017 Jan;5(1):10-22 PMID: 27860400
  48. Acrosome markers of human sperm.
    Anat Sci Int. 2016 Mar;91(2):128-42 PMID: 26748928
  49. The mouse T complex gene Tsga2, encoding polypeptides located in the sperm tail and anterior acrosome, maps to a locus associated with sperm motility and sperm-egg interaction abnormalities.
    Biol Reprod. 2006 Apr;74(4):633-43 PMID: 16354795
  50. The human sperm acrosome reaction: physiology and regulatory mechanisms. An update.
    Hum Reprod Update. 1995 Jan;1(1):51-62 PMID: 9080206
  51. Acrosome Reaction as a Preparation for Gamete Fusion.
    Adv Anat Embryol Cell Biol. 2016;220:159-72 PMID: 27194355
  52. Proteomic characterization of human sperm plasma membrane-associated proteins and their role in capacitation.
    Andrology. 2019 Apr 19;:null PMID: 31002753
  53. New insights into epididymal function in relation to sperm maturation.
    Reproduction. 2013 Dec 19;147(2):R27-42 PMID: 24218627
  54. Analysis of proteomic changes associated with sperm capacitation through the combined use of IPG-strip pre-fractionation followed by RP chromatography LC-MS/MS analysis.
    Proteomics. 2010 Feb;10(3):482-95 PMID: 19943266
  55. Phosphoproteome analysis of capacitated human sperm. Evidence of tyrosine phosphorylation of a kinase-anchoring protein 3 and valosin-containing protein/p97 during capacitation.
    J Biol Chem. 2003 Mar 28;278(13):11579-89 PMID: 12509440
  56. The PRIDE database and related tools and resources in 2019: improving support for quantification data.
    Nucleic Acids Res. 2019 Jan 8;47(D1):D442-D450 PMID: 30395289
  57. Fertility-related proteomic profiling bull spermatozoa separated by percoll.
    J Proteome Res. 2012 Aug 3;11(8):4162-8 PMID: 22794312
  58. A Genome-Wide Haploid Genetic Screen Identifies Heparan Sulfate-Associated Genes and the Macropinocytosis Modulator TMED10 as Factors Supporting Vaccinia Virus Infection.
    J Virol. 2019 Jun 14;93(13): PMID: 30996093
  59. Functional human sperm capacitation requires both bicarbonate-dependent PKA activation and down-regulation of Ser/Thr phosphatases by Src family kinases.
    Mol Hum Reprod. 2013 Sep;19(9):570-80 PMID: 23630234
  60. Identification of capacitation associated tyrosine phosphoproteins in buffalo (Bubalus bubalis) and cattle spermatozoa.
    Anim Reprod Sci. 2011 Jan;123(1-2):40-7 PMID: 21177049
  61. Lysine acetylation modulates mouse sperm capacitation.
    Sci Rep. 2018 Sep 6;8(1):13334 PMID: 30190490
  62. Tssk4 is essential for maintaining the structural integrity of sperm flagellum.
    Mol Hum Reprod. 2015 Feb;21(2):136-45 PMID: 25361759
  63. Evidence for the involvement of proline-rich tyrosine kinase 2 in tyrosine phosphorylation downstream of protein kinase A activation during human sperm capacitation.
    Mol Hum Reprod. 2014 Nov;20(11):1054-66 PMID: 25180269
  64. Mechanism of sperm capacitation and the acrosome reaction: role of protein kinases.
    Asian J Androl. 2012 Nov;14(6):816-21 PMID: 23001443
  65. Sperm Capacitation and Acrosome Reaction in Mammalian Sperm.
    Adv Anat Embryol Cell Biol. 2016;220:93-106 PMID: 27194351
  66. Comparative proteomic identification buffalo spermatozoa during in vitro capacitation.
    Theriogenology. 2019 Mar 1;126:303-309 PMID: 30599421
  67. Acetylproteomic analysis reveals functional implications of lysine acetylation in human spermatozoa (sperm).
    Mol Cell Proteomics. 2015 Apr;14(4):1009-23 PMID: 25680958
  68. Semen proteomics and male infertility.
    J Proteomics. 2017 Jun 6;162:125-134 PMID: 27576136
  69. Proteolytic reaction of mammalian spermatozoa on gelatin membranes.
    Science. 1970 Nov 13;170(3959):749-51 PMID: 4920951
  70. Phosphoglycerate kinase 2 (PGK2) is essential for sperm function and male fertility in mice.
    Biol Reprod. 2010 Jan;82(1):136-45 PMID: 19759366
  71. Involvement of multimeric protein complexes in mediating the capacitation-dependent binding of human spermatozoa to homologous zonae pellucidae.
    Dev Biol. 2011 Aug 15;356(2):460-74 PMID: 21672535
  72. Hyperactivation of capacitated human sperm correlates with the zona pellucida-induced acrosome reaction of zona pellucida-bound sperm.
    Hum Reprod. 2007 Oct;22(10):2632-8 PMID: 17656416
  73. Human sperm activation during capacitation and acrosome reaction: role of calcium, protein phosphorylation and lipid remodelling pathways.
    Front Biosci. 1996 Aug 15;1:d189-205 PMID: 9159227
  74. Heads or tails? Structural events and molecular mechanisms that promote mammalian sperm acrosomal exocytosis and motility.
    Mol Reprod Dev. 2012 Jan;79(1):4-18 PMID: 22031228
  75. Ion channels, phosphorylation and mammalian sperm capacitation.
    Asian J Androl. 2011 May;13(3):395-405 PMID: 21540868
  76. The molecular basis of sperm capacitation.
    J Androl. 1998 Mar-Apr;19(2):242-8 PMID: 9570749
  77. High-throughput sperm differential proteomics suggests that epigenetic alterations contribute to failed assisted reproduction.
    Hum Reprod. 2014 Jun;29(6):1225-37 PMID: 24781426
  78. A comprehensive proteomic approach to identifying capacitation related proteins in boar spermatozoa.
    BMC Genomics. 2014 Oct 14;15:897 PMID: 25315394
  79. Fertilization potential in vitro is correlated with head-specific mannose-ligand receptor expression, acrosome status and membrane cholesterol content.
    Hum Reprod. 1993 Dec;8(12):2155-66 PMID: 8150918
  80. Heat shock protein 90 has roles in intracellular calcium homeostasis, protein tyrosine phosphorylation regulation, and progesterone-responsive sperm function in human sperm.
    PLoS One. 2014 Dec 26;9(12):e115841 PMID: 25541943
  81. Relationship between the zona pellucida (ZP) and ionophore A23187-induced acrosome reaction and the ability of sperm to penetrate the ZP in men with normal sperm-ZP binding.
    Fertil Steril. 1996 Aug;66(2):312-5 PMID: 8690122
  82. The role of the acrosomal matrix in fertilization.
    Int J Dev Biol. 2008;52(5-6):511-22 PMID: 18649264
  83. Protein profile of capacitated versus ejaculated human sperm.
    J Proteome Res. 2009 Jul;8(7):3377-89 PMID: 19408963
  84. Novel signaling pathways involved in sperm acquisition of fertilizing capacity.
    J Reprod Immunol. 2002 Jan;53(1-2):133-50 PMID: 11730911
  85. Sperm head membrane reorganisation during capacitation.
    Int J Dev Biol. 2008;52(5-6):473-80 PMID: 18649260
  86. Methods for the assessment of sperm capacitation and acrosome reaction excluding the sperm penetration assay.
    Methods Mol Biol. 2013;927:113-8 PMID: 22992908
  87. Designing 3-Dimensional In Vitro Oviduct Culture Systems to Study Mammalian Fertilization and Embryo Production.
    Ann Biomed Eng. 2017 Jul;45(7):1731-1744 PMID: 27844174
  88. Quantitative phosphoproteomics analysis reveals a key role of insulin growth factor 1 receptor (IGF1R) tyrosine kinase in human sperm capacitation.
    Mol Cell Proteomics. 2015 Apr;14(4):1104-12 PMID: 25693802
  89. The contribution of human sperm proteins to the development and epigenome of the preimplantation embryo.
    Hum Reprod Update. 2018 Sep 1;24(5):535-555 PMID: 29800303
  90. Biological basis for human capacitation.
    Hum Reprod Update. 2005 May-Jun;11(3):205-14 PMID: 15817522
Article Info
Journal
Frontiers in cell and developmental biology
Abbr.
Front Cell Dev Biol
ISSN
2296-634X
DOI
10.3389/fcell.2019.00295
Published
2019-00-00
Epub
2019-00-20
Pages
295
Language
English
Region
Switzerland
NLM ID
101630250
PMCID
PMC6879431
External Links
PubMed source DOI: 10.3389/fcell.2019.00295 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: [email protected]

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2