Head and flagella subcompartmental proteomic analysis of human spermatozoa

Subcellular proteomics not only deepens our knowledge of what proteins are present within cells, but also opens our understanding as to where those proteins reside. Given the highly differentiated, cross‐linked state of spermatozoa, such studies have proven difficult to perform. In this study we have fractionated spermatozoa into two components, consisting of either the head or flagellar region. Following SDS‐PAGE, 1 mm slices were digested and used for LC‐MS/MS analysis. In total, 1429 proteins were identified with 721 proteins being exclusively found in the tail and 521 exclusively in the head. Not only is this the largest reported proteomic analysis of human spermatozoa, but also it has provided novel insights into the compartmentalization of proteins, particularly receptors, never previously reported to be present in this cell type.

[1]  S. Narumiya,et al.  Identification of Sperm-specific Proteins That Interact with A-kinase Anchoring Proteins in a Manner Similar to the Type II Regulatory Subunit of PKA* , 2001, The Journal of Biological Chemistry.

[2]  A. Kaya,et al.  Delivering value from sperm proteomics for fertility , 2012, Cell and Tissue Research.

[3]  Roy Jones,et al.  Expression and secretion of rat SPAM1(2B1 or PH-20) in the epididymis: role of testicular lumicrine factors. , 2004, Matrix biology : journal of the International Society for Matrix Biology.

[4]  G. McKnight,et al.  The essential role of RI alpha in the maintenance of regulated PKA activity. , 2002, Annals of the New York Academy of Sciences.

[5]  G. McKnight,et al.  The Essential Role of RIα in the Maintenance of Regulated PKA Activity , 2002 .

[6]  R. Aitken,et al.  Reactive oxygen species and human spermatozoa: Analysis of the cellular mechanisms involved in luminol‐ and lucigenin‐dependent chemiluminescence , 1992, Journal of cellular physiology.

[7]  R. Aitken,et al.  Oxidative stress, sperm survival and fertility control , 2006, Molecular and Cellular Endocrinology.

[8]  Marc G Caron,et al.  Dopaminergic Supersensitivity in G Protein-Coupled Receptor Kinase 6-Deficient Mice , 2003, Neuron.

[9]  A. Salicioni,et al.  Signalling pathways involved in sperm capacitation. , 2007, Society of Reproduction and Fertility supplement.

[10]  B. Gadella,et al.  A possible role for sperm RNA in early embryo development. , 2007, Theriogenology.

[11]  C. Scarlett,et al.  Actin polymerisation during morphogenesis of the acrosome as spermatozoa undergo epididymal maturation in the tammar wallaby (Macropus eugenii) , 2001, Journal of anatomy.

[12]  F. Hartl,et al.  Mitochondrial protein import: Identification of processing peptidase and of PEP, a processing enhancing protein , 1988, Cell.

[13]  L. Hall,et al.  Cloning and sequence analysis of rat fertilin alpha and beta--developmental expression, processing and immunolocalization. , 1997, Molecular human reproduction.

[14]  P. Leclerc,et al.  Regulation of the Human Sperm Tyrosine Kinase c-yes. Activation by Cyclic Adenosine 3′,5′-Monophosphate and Inhibition by Ca2+ 1 , 2002, Biology of reproduction.

[15]  G. Gerton,et al.  Identification and validation of mouse sperm proteins correlated with epididymal maturation , 2011, Proteomics.

[16]  J. Nyengaard,et al.  Roles for the pro-neurotrophin receptor sortilin in neuronal development, aging and brain injury , 2007, Nature Neuroscience.

[17]  A. I. Yudin,et al.  The PH-20 protein in human spermatozoa. , 1997, Journal of andrology.

[18]  H. Nojima,et al.  Identification and Characterization of a Haploid Germ Cell-specific Nuclear ProteinKinase (Haspin) in Spermatid Nuclei and Its Effects on Somatic Cells* , 1999, The Journal of Biological Chemistry.

[19]  R. Harrison.  Rapid PKA‐catalysed phosphorylation of boar sperm proteins induced by the capacitating agent bicarbonate , 2004, Molecular reproduction and development.

[20]  A. Sanjay,et al.  Inhibition of Ser/Thr Phosphatases Induces Capacitation-associated Signaling in the Presence of Src Kinase Inhibitors* , 2010, The Journal of Biological Chemistry.

[21]  K. Wüthrich,et al.  NMR structure of the J-domain and the Gly/Phe-rich region of the Escherichia coli DnaJ chaperone. , 1996, Journal of molecular biology.

[22]  J. Rutllant,et al.  Posttranslational Processing of PH-20 During Epididymal Sperm Maturation in the Horse1 , 2001, Biology of reproduction.

[23]  G. Cornwall,et al.  Isolation and Proteomic Characterization of the Mouse Sperm Acrosomal Matrix* , 2012, Molecular & Cellular Proteomics.

[24]  W. Franke,et al.  Molecular nature of calicin, a major basic protein of the mammalian sperm head cytoskeleton. , 1995, Experimental cell research.

[25]  R. Zeng,et al.  Proteomic profiling of regionalized proteins in rat epididymis indicates consistency between specialized distribution and protein functions. , 2006, Journal of proteome research.

[26]  B. N. Day,et al.  Proteasomal Interference Prevents Zona Pellucida Penetration and Fertilization in Mammals1 , 2004, Biology of reproduction.

[27]  P. Stork,et al.  PKA phosphorylation of Src mediates cAMP's inhibition of cell growth via Rap1. , 2002, Molecular cell.

[28]  S. Landry.  Structure and energetics of an allele-specific genetic interaction between dnaJ and dnaK: correlation of nuclear magnetic resonance chemical shift perturbations in the J-domain of Hsp40/DnaJ with binding affinity for the ATPase domain of Hsp70/DnaK. , 2003, Biochemistry.

[29]  W. Willis,et al.  Glyceraldehyde 3-phosphate dehydrogenase-S, a sperm-specific glycolytic enzyme, is required for sperm motility and male fertility. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[30]  M. Baker.  The 'omics revolution and our understanding of sperm cell biology. , 2011, Asian journal of andrology.

[31]  R. Aitken.  The Amoroso Lecture. The human spermatozoon--a cell in crisis? , 1999, Journal of reproduction and fertility.

[32]  G. Daum,et al.  Import of proteins into mitochondria. Partial purification of a matrix-located protease involved in cleavage of mitochondrial precursor polypeptides. , 1983, The Journal of biological chemistry.

[33]  R. Aitken,et al.  Phosphorylation and consequent stimulation of the tyrosine kinase c-Abl by PKA in mouse spermatozoa; its implications during capacitation. , 2009, Developmental biology.

[34]  Rafael Oliva,et al.  Proteomic identification of human sperm proteins , 2006, Proteomics.

[35]  P. Visconti,et al.  Expression analysis of the human testis-specific serine/threonine kinase (TSSK) homologues. A TSSK member is present in the equatorial segment of human sperm. , 2004, Molecular human reproduction.

[36]  H. Galantino-Homer,et al.  Cholesterol Efflux-mediated Signal Transduction in Mammalian Sperm , 1999, The Journal of Biological Chemistry.

[37]  R. Aitken,et al.  Label-free quantitation of phosphopeptide changes during rat sperm capacitation. , 2010, Journal of proteome research.

[38]  P. Primakoff,et al.  Testase 1 (ADAM 24) a plasma membrane-anchored sperm protease implicated in sperm function during epididymal maturation or fertilization. , 2001, Journal of cell science.

[39]  H. Fisher,et al.  Reactive oxygen species generation and human spermatozoa: The balance of benefit and risk , 1994, BioEssays : news and reviews in molecular, cellular and developmental biology.

[40]  P. Cuasnicú,et al.  Peritoneal fluid modifies the response of human spermatozoa to follicular fluid. , 2012, Reproductive biomedicine online.

[41]  D. Koppel,et al.  Lateral diffusion of the PH-20 protein on guinea pig sperm: evidence that barriers to diffusion maintain plasma membrane domains in mammalian sperm , 1987, The Journal of cell biology.

[42]  T. Willnow,et al.  p75NTR – live or let die , 2005, Current Opinion in Neurobiology.

[43]  P. Sutovsky,et al.  Mechanism of sperm-zona pellucida penetration during mammalian fertilization: 26S proteasome as a candidate egg coat lysin. , 2007, Society of Reproduction and Fertility supplement.

[44]  R. Aitken,et al.  Identification of post‐translational modifications that occur during sperm maturation using difference in two‐dimensional gel electrophoresis , 2005, Proteomics.

[45]  T. Willnow,et al.  VPS10P-domain receptors — regulators of neuronal viability and function , 2008, Nature Reviews Neuroscience.

[46]  S. de Mateo,et al.  Proteomic characterization of the human sperm nucleus , 2011, Proteomics.

[47]  F. A. Lai,et al.  PLC zeta: a sperm-specific trigger of Ca(2+) oscillations in eggs and embryo development. , 2002, Development.

[48]  D. Mortimer,et al.  Specific labelling by peanut agglutinin of the outer acrosomal membrane of the human spermatozoon. , 1987, Journal of reproduction and fertility.

[49]  R. Aitken,et al.  The importance of redox regulated pathways in sperm cell biology , 2004, Molecular and Cellular Endocrinology.

[50]  T. Rapoport,et al.  Signal Sequence Processing in Rough Microsomes (*) , 1995, The Journal of Biological Chemistry.

[51]  R. Aitken,et al.  Proteomic insights into the maturation and capacitation of mammalian spermatozoa , 2012, Systems biology in reproductive medicine.

[52]  S. Komatsu,et al.  Fine resolution of human sperm nucleoproteins by two-dimensional electrophoresis. , 2005, Molecular human reproduction.

[53]  N. Cross,et al.  Role of cholesterol in sperm capacitation. , 1998, Biology of reproduction.

[54]  R. Aitken,et al.  The rat sperm proteome characterized via IPG strip prefractionation and LC‐MS/MS identification , 2008, Proteomics.

[55]  S. de Mateo,et al.  Marked correlations in protein expression identified by proteomic analysis of human spermatozoa , 2007, Proteomics.

[56]  L. Hall,et al.  The MDC family of proteins and their processing during epididymal transit. , 1998, Journal of reproduction and fertility. Supplement.

[57]  J. Yates,et al.  A model for random sampling and estimation of relative protein abundance in shotgun proteomics. , 2004, Analytical chemistry.

[58]  R. Aitken,et al.  Identification of SRC as a key PKA-stimulated tyrosine kinase involved in the capacitation-associated hyperactivation of murine spermatozoa , 2006, Journal of Cell Science.

[59]  R. Aitken,et al.  The mouse sperm proteome characterized via IPG strip prefractionation and LC‐MS/MS identification , 2008, Proteomics.

[60]  R. Aitken,et al.  Composition and significance of detergent resistant membranes in mouse spermatozoa , 2009, Journal of cellular physiology.

[61]  S. de Mateo,et al.  Sperm cell proteomics , 2009, Proteomics.

[62]  R. Idzerda,et al.  Mutation of the Calpha subunit of PKA leads to growth retardation and sperm dysfunction. , 2002, Molecular endocrinology.

[63]  T. Muramatsu,et al.  Behaviour of a sperm surface transmembrane glycoprotein basigin during epididymal maturation and its role in fertilization in mice. , 2002, Reproduction.

[64]  W K Whitten,et al.  The culture of mouse embryos in vitro. , 1971 .

[65]  G. Gerton,et al.  Proteomic Profiling of Accessory Structures from the Mouse Sperm Flagellum*S , 2006, Molecular & Cellular Proteomics.

[66]  G. McKnight,et al.  Sperm-specific protein kinase A catalytic subunit Calpha2 orchestrates cAMP signaling for male fertility. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[67]  J. Muschietti,et al.  Bicarbonate dependence of cAMP accumulation induced by phorbol esters in hamster spermatozoa. , 1990, Biochimica et biophysica acta.

[68]  Yan Peng,et al.  c.822+126T>G/C: a novel triallelic polymorphism of the TSSK6 gene associated with spermatogenic impairment in a Chinese population. , 2010, Asian journal of andrology.

[69]  M. Handel,et al.  Protein synthesis in mouse spermatozoa. , 1979, Biology of reproduction.

[70]  D. Edwards,et al.  Human sperm plasma membrane progesterone receptor(s) and the acrosome reaction. , 1996, Biology of reproduction.

[71]  R. Aitken,et al.  Prolactin exerts a prosurvival effect on human spermatozoa via mechanisms that involve the stimulation of Akt phosphorylation and suppression of caspase activation and capacitation. , 2010, Endocrinology.

[72]  L. Mayorga,et al.  Cholesterol content regulates acrosomal exocytosis by enhancing Rab3A plasma membrane association. , 2005, Developmental biology.

[73]  C. Paweletz,et al.  Identification and Characterization of SSTK, a Serine/Threonine Protein Kinase Essential for Male Fertility , 2022 .

[74]  S. Robbins,et al.  Erlin-1 and erlin-2 are novel members of the prohibitin family of proteins that define lipid-raft-like domains of the ER , 2006, Journal of Cell Science.

[75]  R. Lefkowitz,et al.  G protein-coupled receptor kinases. , 1998, Annual review of biochemistry.

[76]  A. Sanjay,et al.  cSrc is necessary for epididymal development and is incorporated into sperm during epididymal transit. , 2012, Developmental biology.

[77]  J. Fujii,et al.  Possible involvement of the membrane-bound form of peroxiredoxin 4 in acrosome formation during spermiogenesis of rats. , 2001, European journal of biochemistry.

[78]  U. Kaupp,et al.  The CatSper channel: a polymodal chemosensor in human sperm , 2012, The EMBO journal.

[79]  P. Sutovsky.  Sperm proteasome and fertilization. , 2011, Reproduction.

[80]  R. Aitken,et al.  The biological significance of detergent-resistant membranes in spermatozoa. , 2009, Journal of reproductive immunology.

[81]  C. Pineau,et al.  Spermatogenesis in mammals: proteomic insights , 2012, Systems biology in reproductive medicine.

[82]  H. Breitbart.  Signaling pathways in sperm capacitation and acrosome reaction. , 2003, Cellular and molecular biology.

[83]  G. Blatch,et al.  Not all J domains are created equal: Implications for the specificity of Hsp40–Hsp70 interactions , 2005, Protein science : a publication of the Protein Society.

[84]  D. Conway,et al.  Population study of causes, treatment, and outcome of infertility. , 1985, British medical journal.

[85]  Stijn Hoorens,et al.  The costs and consequences of assisted reproductive technology: an economic perspective. , 2010, Human reproduction update.

[86]  Y. Gur,et al.  Mammalian sperm translate nuclear-encoded proteins by mitochondrial-type ribosomes. , 2006, Genes & development.

[87]  R. Aitken,et al.  Investigation of the role of SRC in capacitation-associated tyrosine phosphorylation of human spermatozoa. , 2008, Molecular human reproduction.

[88]  M. Baker,et al.  Ontogeny of Tyrosine Phosphorylation-Signaling Pathways During Spermatogenesis and Epididymal Maturation in the Mouse1 , 2006, Biology of reproduction.

[89]  R. Aitken,et al.  DNA Damage in Human Spermatozoa Is Highly Correlated with the Efficiency of Chromatin Remodeling and the Formation of 8-Hydroxy-2′-Deoxyguanosine, a Marker of Oxidative Stress1 , 2009, Biology of reproduction.

[90]  Rakesh Kumar,et al.  Transforming growth factor β1 enhances expression of 50 kDa protein related to 2′‐5′ oligoadenylate synthetase in human sperm cells , 1991 .

[91]  R. Aitken,et al.  Analysis of the ability of three probes targeting the outer acrosomal membrane or acrosomal contents to detect the acrosome reaction in human spermatozoa. , 1993, Human reproduction.

[92]  J. Garin,et al.  Proteomic strategy for the identification of critical actors in reorganization of the post-meiotic male genome. , 2012, Molecular human reproduction.

[93]  R. Aitken,et al.  Identification of gene products present in Triton X‐100 soluble and insoluble fractions of human spermatozoa lysates using LC‐MS/MS analysis , 2007, Proteomics. Clinical applications.

[94]  R. Aitken,et al.  The role of proteomics in understanding sperm cell biology. , 2008, International journal of andrology.

[95]  R. Aitken.  Possible redox regulation of sperm motility activation. , 2000, Journal of andrology.

[96]  N. A. Spiridonov,et al.  Tssk6 is required for Izumo relocalization and gamete fusion in the mouse , 2009, Journal of Cell Science.

[97]  B. A. Lessley,et al.  Isolation of motile spermatozoa by density gradient centrifugation in Percoll , 1983 .

[98]  G. Kopf,et al.  Capacitation of mouse spermatozoa. I. Correlation between the capacitation state and protein tyrosine phosphorylation. , 1995, Development.

[99]  Barbara L. Hempstead,et al.  Sortilin is essential for proNGF-induced neuronal cell death , 2004, Nature.

[100]  V. De Pinto,et al.  VDAC1 Is a Transplasma Membrane NADH-Ferricyanide Reductase* , 2004, Journal of Biological Chemistry.

[101]  William Arbuthnot Sir Lane,et al.  Proteomic analysis of sperm regions that mediate sperm‐egg interactions , 2006, Proteomics.

[102]  G S Kopf,et al.  Capacitation of mouse spermatozoa. II. Protein tyrosine phosphorylation and capacitation are regulated by a cAMP-dependent pathway. , 1995, Development.

[103]  P. Sutovsky,et al.  The sperm proteasome during sperm capacitation and fertilization. , 2009, Journal of reproductive immunology.

[104]  J. Jiwakanon,et al.  Short communication: Concentration of TGF-β1, IL-10 and IL-6 in boar seminal plasma and TGF-β1 level in different fractions of ejaculates. , 2012, Animal reproduction science.