Complementarity between sperm surface β-l,4-galactosyl-transferase and egg-coat ZP3 mediates sperm–egg binding

Nature ◽  
1992 ◽  
Vol 357 (6379) ◽  
pp. 589-593 ◽  
Author(s):  
David J. Miller ◽  
Mary B. Macek ◽  
Barry D. Shur
Keyword(s):  
Sperm Surface ◽  
Galactosyl Transferase ◽  
Egg Coat

Sperm from beta 1,4-galactosyltransferase-null mice are refractory to ZP3-induced acrosome reactions and penetrate the zona pellucida poorly

Development ◽  
1997 ◽  
Vol 124 (20) ◽  
pp. 4121-4131 ◽  
Author(s):  
Q. Lu ◽  
B.D. Shur
Keyword(s):  
Acrosome Reaction ◽  
Direct Evidence ◽  
Calcium Ionophore ◽  
Wild Type ◽  
Sperm Surface ◽  
Gamete Recognition ◽  
Dependent Signal ◽  
Relative Inability ◽  
Egg Coat

A variety of sperm surface components have been suggested to mediate gamete recognition by binding to glycoside ligands on the egg coat glycoprotein ZP3. The function of each of these candidate receptors is based upon varying degrees of circumstantial and direct evidence; however, the effects on fertilization of targeted mutations in any of these candidate receptors have not yet been reported. In this paper, we describe the effects of targeted mutations in beta1,4-galactosyltransferase, the best studied of the candidate receptors for ZP3. Surprisingly, galactosyltransferase-null (gt[−/−]) males are fertile; however, sperm from gt(−/−) males bind less radiolabeled ZP3 than wild-type sperm, and are unable to undergo the acrosome reaction in response to either ZP3 or anti-galactosyltransferase antibodies, as do wild-type sperm. In contrast, gt(−/−) sperm undergo the acrosome reaction normally in response to calcium ionophore, which bypasses the requirement for ZP3 binding. The inability of gt(−/−) sperm to undergo a ZP3-induced acrosome reaction renders them physiologically inferior to wild-type sperm, as assayed by their relative inability to penetrate the egg coat and fertilize the oocyte in vitro. Thus, although ZP3 binding and subsequent induction of the acrosome reaction are dispensable for fertilization, they impart a physiological advantage to the fertilizing sperm. A second strain of mice was created that is characterized by a loss of of the long galactosyltransferase isoform responsible for ZP3-dependent signal transduction, but which maintains normal levels of Golgi galactosylation. Sperm from these mice show that the defective sperm-egg interactions in gt(−/−) mice are due directly to a loss of the long galactosyltransferase isoform from the sperm surface and are independent of the state of intracellular galactosylation during spermatogenesis.

Spermatozoa from a marsupial, the brushtail possum, contain β1,4-galactosyltransferase

2008 ◽  
Vol 20 (3) ◽  
pp. 402 ◽  
Author(s):  
A. G. Braundmeier ◽  
William G. Breed ◽  
D. J. Miller
Keyword(s):  
Molecular Weight ◽  
Zona Pellucida ◽  
Acrosome Reaction ◽  
Brushtail Possum ◽  
Laboratory Mice ◽  
Sperm Surface ◽  
Protein Functions ◽  
Sperm Acrosome Reaction ◽  
Porcine Spermatozoa ◽  
Egg Coat

β1,4-Galactosyltransferase-I (GalTase-I) is one of the key molecules on the sperm surface of eutherian mammals that is likely to be involved in binding to the egg coat, the zona pellucida, to mediate sperm–egg interaction. In laboratory mice, the species for which most data are available, this protein functions as a receptor for the zona pellucida protein ZP3 of the oocyte and, upon binding, triggers the sperm acrosome reaction. In the present study, we investigated the presence and abundance of GalTase-I in epididymal sperm extracts of a marsupial, the brushtail possum, Trichosurus vulpecula. For this, spermatozoa were collected from cauda epididymides and the amount of β1,4-galactosyltransferase activity in washed sperm extracts was compared with that of porcine spermatozoa. Overall β1,4-galactosyltransferase enzyme activity was found to be more abundant in possum sperm extracts than those from porcine spermatozoa (P < 0.05). Immunoblots with an antibody to mouse GalTase-I revealed that the molecular weight of possum spermatozoa GalTase-I was 66 kDa, which is similar to the molecular weight of GalTase-I in spermatozoa from eutherian mammals. The molecular weight of GalTase-I was the same in sperm extracts collected from the caput and cauda epididymides. These results demonstrate that GalTase-I is indeed present in possum spermatozoa and thus it may be a gamete receptor molecule on the sperm surface of marsupials as well as those of eutherian mammals.

scholarly journals A role for mouse sperm surface galactosyltransferase in sperm binding to the egg zona pellucida.

1982 ◽  
Vol 95 (2) ◽  
pp. 574-579 ◽  
Author(s):  
B D Shur ◽  
N G Hall
Keyword(s):  
Zona Pellucida ◽  
Milk Protein ◽  
Preceding Paper ◽  
Transferase Activity ◽  
Mouse Sperm ◽  
Sperm Surface ◽  
Galactosyl Transferase ◽  
Sperm Binding ◽  
Beta Galactosidase

Past studies have suggested that mouse sperm surface galactosyltransferase may participate during fertilization by binding N-acetylglucosamine (GlcNAc) residues in the zona pellucida. In this paper, we examined further the role of sperm surface galactosyltransferase in mouse fertilization. Two reagents that specifically perturb sperm surface galactosyltransferase activity both inhibit sperm-zona binding. The presence of the milk protein alpha-lactalbumin specifically modifies the substrate specificity of sperm galactosyltransferase away from GlcNAc and towards glucose and simultaneously inhibits sperm binding to the zona pellucida. Similarly, UDP-dialdehyde inhibits sperm binding to the zona pellucida and sperm surface galactosyl-transferase activity to identical degrees. Of five other sperm enzymes assayed, four are unaffected by UDP-dialdehyde, and one is affected only slightly. Covalent linkage of UDP-dialdehyde to sperm dramatically inhibits binding to eggs, while treatment of eggs with UDP-dialdehyde has no effect on sperm binding. Heat-solubilized or pronase-digested zona pellucida inhibit sperm-zona binding, and they can be glycosylated by sperm with UDP-galactose. Sperm are also able to glycosylate intact zona pellucida with UDP-galactose. Thus, solubilized and intact zona pellucida act as substrates for sperm surface GlcNAc:galactosyltransferases. Finally, pretreatment of eggs with beta-N-acetylglucosaminidase inhibits sperm binding by up to 86%, while under identical conditions, pretreatment with beta-galactosidase increases sperm binding by 55%. These studies, in conjunction with those of the preceding paper dealing with surface galactosyltransferase changes during capacitation, directly suggest that galactosyltransferase is at least one of the components necessary for sperm binding to the zona pellucida.

scholarly journals Overexpressing sperm surface beta 1,4-galactosyltransferase in transgenic mice affects multiple aspects of sperm-egg interactions.

1994 ◽  
Vol 126 (6) ◽  
pp. 1573-1583 ◽  
Author(s):  
A Youakim ◽  
H J Hathaway ◽  
D J Miller ◽  
X Gong ◽  
B D Shur
Keyword(s):  
Transgenic Mice ◽  
Zona Pellucida ◽  
Transgenic Animals ◽  
Wild Type ◽  
Sperm Surface ◽  
Receptor Aggregation ◽  
Excess Surface ◽  
Before And After ◽  
Mechanistic Basis ◽  
Egg Coat

Sperm surface beta 1,4-galactosyltransferase (GalTase) mediates fertilization in mice by binding to specific O-linked oligosaccharide ligands on the egg coat glycoprotein ZP3. Before binding the egg, sperm GalTase is masked by epididymally derived glycosides that are shed from the sperm surface during capacitation. After binding the egg, sperm-bound oligosaccharides on ZP3 induce the acrosome reaction by receptor aggregation, presumably involving GalTase. In this study, we asked how increasing the levels of sperm surface GalTase would affect sperm-egg interactions using transgenic mice that overexpress GalTase under the control of a heterologous promoter. GalTase expression was elevated in many tissues in adult transgenic animals, including testis. Sperm from transgenic males had approximately six times the wild-type level of surface GalTase protein, which was localized appropriately on the sperm head as revealed by indirect immunofluorescence. As expected, sperm from transgenic mice bound more radiolabeled ZP3 than did wild-type sperm. However, sperm from transgenic animals were relatively unable to bind eggs, as compared to sperm from wild-type animals. The mechanistic basis for the reduced egg-binding ability of transgenic sperm was attributed to alterations in two GalTase-dependent events. First, transgenic sperm that overexpress surface GalTase bound more epididymal glycoside substrates than did sperm from wild-type mice, thus masking GalTase and preventing it from interacting with its zona pellucida ligand. Second, those sperm from transgenic mice that were able to bind the zona pellucida were hypersensitive to ZP3, such that they underwent precocious acrosome reactions and bound to eggs more tenuously than did wild-type sperm. These results demonstrate that sperm-egg binding requires an optimal, rather than maximal, level of surface GalTase expression, since increasing this level decreases sperm reproductive efficiency both before and after egg binding. Although sperm GalTase is required for fertilization by serving as a receptor for the egg zona pellucida, excess surface GalTase is counterproductive to successful sperm-egg binding.

Immunocytochemical localization of β1-4 galactosyltransferase in endometrial carcinoma cells

1990 ◽  
Vol 48 (3) ◽  
pp. 944-945
Author(s):  
Ichiro Yamamoto ◽  
Toshiaki Tachibana ◽  
Hiroko Maruyama ◽  
Noriyuki Komatsu ◽  
Hiroyuki Kuramoto ◽  
...  
Keyword(s):  
Endometrial Carcinoma ◽  
Cell Lines ◽  
Endometrial Adenocarcinoma ◽  
Protein A ◽  
Rabbit Antiserum ◽  
Carcinoma Cells ◽  
Affinity Column ◽  
Antibody Technique ◽  
Galactosyl Transferase ◽  
C Terminus

We have paid attention to the alteration of glycosyltransferase in carcinoma cells, because it might be related to the malignancy of the cells. In this connection, localization of β1-4 galactosyl transferase (β1-4 Gal T) in human endometrial carcinoma cells was examined immunocytochemically using two kinds of cell lines, each of which showed different degree of differentiation.An antibody was purified from the rabbit antiserum against the synthetic peptide, IFNRLVFRGMSC (W89) of human β1-4 Gal T coupled with KLH (keyhole limpet hemocyanine) by protein A column and peptide-affinity column chromatography. The anti-W89 serum reacts to the C-terminus of human β 1-4 Gal T and to both membrane-bound and soluble forms of the enzyme. Cell line of well differentiated endometrial adenocarcinoma (I) and that of poorly differentiated endometrial adenocarcinoma (50B) were cultivated respectively in MEM medium containing 15% FCS and 2 mM glutamine for 4 d at 37°C under 5% CO2. The cells were fixed in a mixture of 4% paraformaldehyde and 0.1% glutaraldehyde in 0.1 M Soerensen’s phosphate buffer (pH 7.4) at 4°C for 30 min, washed with PBS, then freezed and thawed. The indirect method of the peroxidase- labeled antibody technique was used for immunocytochemistry of both LM and TEM on the cell lines. The cells were dehydrated in ethanol and embedded in TAAB 812. Ultrathin sections were observed under a TEM, JEM-100S.

scholarly journals Buffalo sperm surface proteome profiling reveals an intricate relationship between innate immunity and reproduction

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Vipul Batra ◽  
Vanya Bhushan ◽  
Syed Azmal Ali ◽  
Parul Sarwalia ◽  
Ankit Pal ◽  
...  
Keyword(s):  
Male Fertility ◽  
Female Reproductive Tract ◽  
Surface Proteins ◽  
Glycosylation Pattern ◽  
Sperm Surface ◽  
Beta Defensins ◽  
Surface Proteome ◽  
Related Proteins ◽  
Sperm Surface Proteins

Abstract Background Low conception rate (CR) despite insemination with morphologically normal spermatozoa is a common reproductive restraint that limits buffalo productivity. This accounts for a significant loss to the farmers and the dairy industry, especially in agriculture-based economies. The immune-related proteins on the sperm surface are known to regulate fertility by assisting the spermatozoa in their survival and performance in the female reproductive tract (FRT). Regardless of their importance, very few studies have specifically catalogued the buffalo sperm surface proteome. The study was designed to determine the identity of sperm surface proteins and to ascertain if the epididymal expressed beta-defensins (BDs), implicated in male fertility, are translated and applied onto buffalo sperm surface along with other immune-related proteins. Results The raw mass spectra data searched against an in-house generated proteome database from UniProt using Comet search engine identified more than 300 proteins on the ejaculated buffalo sperm surface which were bound either by non-covalent (ionic) interactions or by a glycosylphosphatidylinositol (GPI) anchor. The singular enrichment analysis (SEA) revealed that most of these proteins were extracellular with varied binding activities and were involved in either immune or reproductive processes. Flow cytometry using six FITC-labelled lectins confirmed the prediction of glycosylation of these proteins. Several beta-defensins (BDs), the anti-microbial peptides including the BuBD-129 and 126 were also identified amongst other buffalo sperm surface proteins. The presence of these proteins was subsequently confirmed by RT-qPCR, immunofluorescence and in vitro fertilization (IVF) experiments. Conclusions The surface of the buffalo spermatozoa is heavily glycosylated because of the epididymal secreted (glyco) proteins like BDs and the GPI-anchored proteins (GPI-APs). The glycosylation pattern of buffalo sperm-surface, however, could be perturbed in the presence of elevated salt concentration or incubation with PI-PLC. The identification of numerous BDs on the sperm surface strengthens our hypothesis that the buffalo BDs (BuBDs) assist the spermatozoa either in their survival or in performance in the FRT. Our results suggest that BuBD-129 is a sperm-surface BD that could have a role in buffalo sperm function. Further studies elucidating its exact physiological function are required to better understand its role in the regulation of male fertility.

Dispersal of sperm surface antigens in the plasma membranes of polyspermically fertilized sea urchin eggs

1987 ◽  
Vol 173 (2) ◽  
pp. 628-632 ◽  
Author(s):  
David Nishioka ◽  
Donald C. Porter ◽  
James S. Trimmer ◽  
Victor D. Vacquier
Keyword(s):  
Sea Urchin ◽  
Plasma Membranes ◽  
Surface Antigens ◽  
Sea Urchin Eggs ◽  
Sperm Surface
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