scholarly journals The sea urchin egg receptor for sperm: isolation and characterization of the intact, biologically active receptor

1993 ◽  
Vol 122 (4) ◽  
pp. 887-895 ◽  
Author(s):  
K Ohlendieck ◽  
ST Dhume ◽  
JS Partin ◽  
WJ Lennarz
Keyword(s):  
Cell Surface ◽  
Sea Urchin ◽  
Transmembrane Protein ◽  
Biologically Active ◽  
Cell Surface Receptor ◽  
Egg Cell ◽  
Dependent Manner ◽  
Binding Studies ◽  
Isolation And Characterization ◽  
Species Specific

The species-specific binding of sea urchin sperm to the egg is mediated by an egg cell surface receptor. Although earlier studies have resulted in the cloning and sequencing of the receptor, structure/function studies require knowledge of the structure of the mature cell surface protein. In this study, we report the purification of this glycoprotein to homogeneity from a cell surface complex of Strongylocentrotus purpuratus eggs using lectin and ion exchange chromatography. Based on the yield of receptor it can be calculated that each egg contains approximately 1.25 x 10(6) receptor molecules on its surface. The receptor, which has an apparent M(r) of 350 kD, is a highly glycosylated transmembrane protein composed of approximately 70% carbohydrate. Because earlier studies on the partially purified receptor and on a pure, extracellular fragment of the receptor indicated that the carbohydrate chains were important in sperm binding, we undertook compositional analysis of the carbohydrate in the intact receptor. These analyses and lectin binding studies revealed that the oligosaccharide chains of the receptor are sulfated and that both N- and O-linked chains are present. Functional analyses revealed that the purified receptor retained biological activity; it inhibited fertilization in a species-specific and dose-dependent manner, and polystyrene beads coated with it bound to acrosome-reacted sperm in a species-specific manner. The availability of biochemical quantities of this novel cell recognition molecule opens new avenues to studying the interaction of complementary cell surface ligands in fertilization.

scholarly journals Identification of the sea urchin egg receptor for sperm using an antiserum raised against a fragment of its extracellular domain.

1992 ◽  
Vol 116 (3) ◽  
pp. 647-658 ◽  
Author(s):  
K R Foltz ◽  
W J Lennarz
Keyword(s):  
Cell Surface ◽  
Sea Urchin ◽  
Egg Cell ◽  
Cortical Granule ◽  
Dependent Manner ◽  
Western Blots ◽  
Cell Surface Glycoprotein ◽  
Sperm Binding ◽  
Sea Urchin Egg ◽  
Species Specific

Sea urchin egg fertilization requires the species-specific interaction of molecules on the sperm and egg surfaces. Previously, we isolated an extracellular, 70-kD glycosylated fragment of the S. purpuratus egg receptor for sperm by treating the eggs with lysylendoproteinase C (Foltz, K. R., and W. J. Lennarz. 1990. J. Cell Biol. 111:2951-2959). To characterize the receptor further, we have generated a polyclonal antiserum (anti-70KL) against the purified 70-kD fragment. Anti-70KL was found to react with a single polypeptide of approximately 350 kD on Western blots, presumed to be the intact receptor, in an egg cell surface preparation. This polypeptide appeared to be tightly associated with the plasma membrane/vitelline layer complex, as it was released from these preparations only by detergent treatment. Immunofluorescence microscopy revealed that the receptor was distributed evenly over the egg surface. The anti-70KL was species specific both in its ability to recognize the egg surface protein and to inhibit sperm binding. Fab fragments generated from affinity-purified anti-70KL also bound to the egg surface and inhibited sperm binding in a concentration-dependent manner. Interestingly, treatment with Fabs caused a small percentage of eggs to undergo cortical granule exocytosis, even in the absence of external Ca2+. These results confirm earlier findings indicating that the receptor is a cell surface glycoprotein of high molecular weight that species specifically binds sperm. This antiserum provides a powerful tool for further investigation of gamete interactions and the structure of the sperm receptor.

scholarly journals Bindin is essential for fertilization in the sea urchin

2021 ◽  
Vol 118 (34) ◽  
pp. e2109636118
Author(s):  
Gary M. Wessel ◽  
Yuuko Wada ◽  
Mamiko Yajima ◽  
Masato Kiyomoto
Keyword(s):  
Cell Surface ◽  
Sea Urchin ◽  
Critical Role ◽  
Egg Cell ◽  
Surface Binding ◽  
Broadcast Spawning ◽  
Cell Surface Binding ◽  
Insoluble Particles ◽  
Males And Females ◽  
Species Specific

Species-specific sperm−egg interactions are essential for sexual reproduction. Broadcast spawning of marine organisms is under particularly stringent conditions, since eggs released into the water column can be exposed to multiple different sperm. Bindin isolated from the sperm acrosome results in insoluble particles that cause homospecific eggs to aggregate, whereas no aggregation occurs with heterospecific eggs. Therefore, Bindin is concluded to play a critical role in fertilization, yet its function has never been tested. Here we report that Cas9-mediated inactivation of the bindin gene in a sea urchin results in perfectly normal-looking embryos, larvae, adults, and gametes in both males and females. What differed between the genotypes was that the bindin−/− sperm never fertilized an egg, functionally validating Bindin as an essential gamete interaction protein at the level of sperm–egg cell surface binding.

Fertilistion in sea urchins: how many different molecules are involved in gamete interaction and fusion?

Zygote ◽  
1994 ◽  
Vol 2 (1) ◽  
pp. 1-4 ◽  
Author(s):  
William J. Lennarz
Keyword(s):  
Cell Surface ◽  
Sea Urchin ◽  
Sea Urchins ◽  
Structure And Function ◽  
Cell Surface Receptor ◽  
Egg Cell ◽  
Cell Surface Molecules ◽  
Surface Molecules ◽  
Surface Receptor ◽  
And Function

SummaryIt has been established that fertilisation in the sea urchin involves binding of acrosome-reacted sperm to an egg cell surface receptor. The structure and function of receptor, as well as the possible involvement of other cell surface molecules in the binding, fusion and activation events, is discussed.

Immunogold and immunoblot studies on a cell surface protein found in primary mesenchyme cells and in the cortical secretory vesicles of the sea urchin egg

1987 ◽  
Vol 45 ◽  
pp. 832-833
Author(s):  
G.L. Decker ◽  
M.C. Valdizan
Keyword(s):  
Cell Surface ◽  
Sea Urchin ◽  
Surface Protein ◽  
Egg Cell ◽  
Secretory Vesicles ◽  
Cell Surface Protein ◽  
Surface Complexes ◽  
Mesenchyme Cells ◽  
Lowicryl K4m ◽  
Primary Mesenchyme Cells

A monoclonal antibody designated MAb 1223 has been used to show that primary mesenchyme cells of the sea urchin embryo express a 130-kDa cell surface protein that may be directly involved in Ca2+ uptake required for growth of skeletal spicules. Other studies from this laboratory have shown that the 1223 antigen, although in relatively low abundance, is also expressed on the cell surfaces of unfertilized eggs and on the majority of blastomeres formed prior to differentiation of the primary mesenchyme cells.We have studied the distribution of 1223 antigen in S. purpuratus eggs and embryos and in isolated egg cell surface complexes that contain the cortical secretory vesicles. Specimens were fixed in 1.0% paraformaldehyde and 1.0% glutaraldehyde and embedded in Lowicryl K4M as previously reported. Colloidal gold (8nm diameter) was prepared by the method of Mulpfordt.

scholarly journals The cell surface receptor for immunoglobulin E. II. Properties of the purified biologically active receptor.

1979 ◽  
Vol 254 (9) ◽  
pp. 3194-3200
Author(s):  
A. Kulczycki ◽  
B L Hempstead ◽  
S L Hofmann ◽  
E W Wood ◽  
C W Parker
Keyword(s):  
Cell Surface ◽  
Immunoglobulin E ◽  
Biologically Active ◽  
Cell Surface Receptor ◽  
Surface Receptor

scholarly journals Dectin-1-Mediated Production of Pro-Inflammatory Cytokines Induced by Yeast β-Glucans in Bovine Monocytes

2021 ◽  
Vol 12 ◽  
Author(s):  
Ana R. V. Pedro ◽  
Tânia Lima ◽  
Ricardo Fróis-Martins ◽  
Bárbara Leal ◽  
Isabel C. Ramos ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Surface ◽  
Inflammatory Cytokines ◽  
Surface Expression ◽  
Cell Surface Receptor ◽  
Dependent Manner ◽  
Feed Supplements ◽  
Surface Receptor ◽  
Dose Dependent ◽  
Pro Inflammatory Cytokines

Yeast-derived products containing β-glucans have long been used as feed supplements in domesticated animals in an attempt to increase immunity. β-glucans are mainly recognized by the cell surface receptor CLEC7A, also designated Dectin-1. Although the immune mechanisms elicited through Dectin-1 activation have been studied in detail in mice and humans, they are poorly understood in other species. Here, we evaluated the response of bovine monocytes to soluble and particulate purified β-glucans, and also to Zymosan. Our results show that particulate, but not soluble β-glucans, can upregulate the surface expression of costimulatory molecules CD80 and CD86 on bovine monocytes. In addition, stimulated cells increased production of IL-8 and of TNF, IL1B, and IL6 mRNA expression, in a dose-dependent manner, which correlated positively with CLEC7A gene expression. Production of IL-8 and TNF expression decreased significantly after CLEC7A knockdown using two different pairs of siRNAs. Overall, we demonstrated here that bovine monocytes respond to particulate β-glucans, through Dectin-1, by increasing the expression of pro-inflammatory cytokines. Our data support further studies in cattle on the induction of trained immunity using dietary β-glucans.

The putative sponge aggregation receptor. Isolation and characterization of a molecule composed of scavenger receptor cysteine-rich domains and short consensus repeats

1998 ◽  
Vol 111 (17) ◽  
pp. 2635-2644 ◽  
Author(s):  
B. Blumbach ◽  
Z. Pancer ◽  
B. Diehl-Seifert ◽  
R. Steffen ◽  
J. Munkner ◽  
...  
Keyword(s):  
Cell Surface ◽  
Scavenger Receptor ◽  
Cell Surface Receptor ◽  
Functional Assay ◽  
Cdna Clones ◽  
The Third ◽  
Short Consensus Repeats ◽  
Isolation And Characterization ◽  
A Cell ◽  
Cell Cell

Porifera (sponges) are the oldest extant metazoan phylum. Dissociated sponge cells serve as a classic system to study processes of cell reaggregation. The reaggregation of dissociated cells is mediated by an extracellularly localized aggregation factor (AF), based on heterophilic interactions of the third order; the AF bridges two cells by ligating a cell-surface-bound aggregation receptor (AR). In the present study we report cloning, expression and immunohistochemical localization of a polypeptide from the marine sponge Geodia cydonium, which very likely represents the AR. The presumed AR gene gives rise to at least three forms of alternatively spliced transcripts of 6.5, 4.9 and 3.9 kb, as detected by northern blotting. Two cDNA clones corresponding to the shorter forms were already reported earlier; here we present an analysis of the largest. All three putative polypeptides feature scavenger receptor cysteine-rich (SRCR) domains. The largest form, SRCR-SCR-Car, is a cell-surface receptor of molecular mass 220 kDa, which is assumed to be the cell-adhesion receptor AR; the second form, SRCR-Re, is also a putative receptor of 166 kDa, while the third form, SRCR-Mo, is a soluble molecule of 129 kDa. The SRCR-SCR-Car molecule consists of fourteen SRCR domains, six short consensus repeats (SCRs), a C-terminal transmembrane domain and a cytoplasmic tail; its fourteenth SRCR domain features an Arg-Gly-Asp tripeptide. To obtain monoclonal antibodies, a 170-amino-acid-long polypeptide that is found in all three forms of the SRCR-containing proteins was expressed in E. coli. In a western blot of sponge cells lysate the monoclonal antibody raised against the recombinant polypeptide recognized two major immuno-reacting polypeptides (220 and 117 kDa) and two minor bands (36 and 32 kDa). The antibody was found to react with antigen(s) predominantly localized on the plasma membranes of cells, especially those of spherulous cells. In a functional assay Fab' fragments of the antibodies suppressed AF-mediated cell-cell reaggregation. Additionally, a recombinant SRCR-soluble fragment effectively inhibited AF-mediated cell-cell reaggregation. We conclude that the 220 kDa SRCR-containing protein of the sponge G. cydonium is very likely the AR.

scholarly journals Yaba-like disease virus protein 7L is a cell-surface receptor for chemokine CCL1

2003 ◽  
Vol 84 (12) ◽  
pp. 3325-3336 ◽  
Author(s):  
Pilar Najarro ◽  
Han-Joo Lee ◽  
James Fox ◽  
James Pease ◽  
Geoffrey L. Smith
Keyword(s):  
Cell Surface ◽  
Disease Virus ◽  
Chemokine Receptor ◽  
Amino Acid Identity ◽  
Cell Surface Receptor ◽  
Cell Protein ◽  
Virus Protein ◽  
Expression Of Genes ◽  
Surface Receptor ◽  
Species Specific

Yaba-like disease virus (YLDV) genes 7L and 145R are located on opposite ends of the genome and are predicted to encode 7-transmembrane proteins (7-TM) that share 53 and 44 % amino acid identity, respectively, to human CC chemokine receptor 8 (hCCR8). In this report, we demonstrate that early after infection with YLDV, cells acquire the ability to bind human CCL1. By expression of genes 7L and 145R in vaccinia virus, we demonstrated that each protein is glycosylated and is exposed on the cell surface with the N terminus outside the cell. Protein 7L, but not 145R, is able to bind hCCL1 (K d=0·6±0·13 nM) and couple to heterotrimeric G-proteins and to activate the extracellular signal-regulated kinases (ERK1/2). 7L binds several chemokines including the viral chemokines vMIPI and vMIPII and hCCL7/MCP3. This binding seems species-specific as 7L does not bind the murine orthologues of CCL1 and CCL7 in the assays used. This represents the first example of a poxviral 7-TM chemokine receptor that has functional interactions with a human chemokine.

scholarly journals Inhibitory Effects of Echinochrome A, Isolated from Shell of the Sea Urchin Anthocidaris crassispina, on Antigen-Stimulated Degranulation in Rat Basophilic Leukemia RBL-2H3 Cells through Suppression of Lyn Activation

2016 ◽  
Vol 11 (9) ◽  
pp. 1934578X1601100
Author(s):  
Tomohiro Itoh ◽  
Azusa Fujiwara ◽  
Masayuki Ninomiya ◽  
Toshimichi Maeda ◽  
Masashi Ando ◽  
...  
Keyword(s):  
Sea Urchin ◽  
Intracellular Signaling ◽  
Immunoglobulin E ◽  
Inhibitory Effect ◽  
Biologically Active ◽  
Type I ◽  
Dependent Manner ◽  
Echinochrome A ◽  
Basophilic Leukemia ◽  
Anthocidaris Crassispina

Echinochrome A (Echi-A) was isolated from the sea urchin Anthocidaris crassispina and its structure determined using 1D and 2D-NMR. In the present study, we examined the inhibitory effect of Echi-A on antigen-stimulated degranulation in rat basophilic leukemia RBL-2H3 cells, which were suppressed in a dose dependent manner. The antigens bind to the high affinity immunoglobulin E receptor, which is expressed on the surface of mast cells and basophils and activate intracellular signal transduction, resulting in the release of biologically active mediators such as histamine. In order to disclose the inhibitory mechanisms of degranulation by Echi-A, we examined the elevation in intracellular Ca2+ concentration ([Ca2+]i), production levels of intracellular reactive oxygen species (ROS) and early intracellular signaling events. Both elevation of [Ca2+]i and intracellular ROS production were markedly suppressed in cells treated with Echi-A. Echi-A also suppressed the activation of Lyn, Syk, and PLCγ1/2 in antigen-stimulated cells. These results indicated that inhibition of antigen-stimulated degranulation in RBL-2H3 cells by Echi-A is mainly due to the inactivation of Lyn/Syk/PLCγ signaling pathways. Our findings suggest that Echi-A could be a beneficial agent for alleviating the symptoms of type I allergy.

Apolipoprotein(a)-Dependent Inhibition of Pericellular Plasminogen Activation Is Mediated by Specific Cellular Receptors

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2236-2236
Author(s):  
Rocco Romagnuolo ◽  
Michael B Boffa ◽  
Marlys L Koschinsky
Keyword(s):  
Cell Surface ◽  
Plasminogen Activator ◽  
Conflicts Of Interest ◽  
Cell Surface Receptor ◽  
Dependent Manner ◽  
Plasminogen Activation ◽  
Apolipoprotein A ◽  
Type Plasminogen Activator ◽  
Urokinase Type Plasminogen Activator

Abstract Abstract 2236 Lipoprotein(a) [Lp(a)] has been identified as an independent risk factor for cardiovascular diseases such as coronary heart disease. Lp(a) levels vary over 1000-fold within the human population and Lp(a) possesses both proatherogenic and prothrombotic properties due to the LDL-like moiety and apolipoprotein(a) [apo(a)] components, respectively. Apo(a) is highly homologous to plasminogen and thus can potentially interfere with plasminogen activation. Plasmin generated in the context of fibrin mediates the breakdown of blood clots, which are the causative factors in heart attacks and strokes. Plasmin generated on the surface of vascular cells plays a role in cell migration and proliferation, two of the fibroproliferative inflammatory events that underlie atherosclerosis. Previous studies have suggested that apo(a) may inhibit pericellular plasminogen activation on the basis of observations that apo(a) decreases plasminogen binding to cells. We have undertaken analysis of the mechanism by which apo(a) may interfere with pericellular plasminogen activation to allow for a more definitive description of the role of Lp(a) within the vasculature. Plasminogen activation was found to be markedly inhibited by the recombinant apo(a) variant 17K, in a dose dependent manner, on human umbilical vein endothelial cells (HUVECs), human monocytic leukemia cells (THP-1), THP-1 macrophages, and smooth muscle cells. The strong lysine binding site in kringle IV type 10, as well as kringle V appear to be required for this effect since apo(a) variants lacking these elements (17KΔAsp and 17KΔV, respectively) failed to inhibit activation. However, the role of lysine-dependent binding of apo(a) itself to the cells is not clear. Carboxypeptidase treatment of cells did not decrease apo(a) binding, and apo(a) does not compete directly for plasminogen binding to the cells. Rather, apo(a) and plasminogen may bind to the cells as a complex. We next attempted to identify the cell-surface receptor(s) that mediate plasminogen activation on the cell surface as well as its inhibition by apo(a). Urokinase-type plasminogen activator receptor (uPAR) has been previously shown to bind to urokinase-type plasminogen activator (uPA), vitronectin, and β3 integrins. uPAR is involved in the remodeling of the extracellular matrix (ECM) through regulation of plasminogen activation. We found evidence that uPAR is a potential receptor for both plasminogen and apo(a). Knockdown of uPAR in HUVECs results in decreased binding of plasminogen, 17K and, to a lesser extent, 17KΔAsp and 17KΔV. Similar experiments in SMCs revealed no changes in binding. A decrease in tPA-mediated plasminogen activation following uPAR knockdown occurred in HUVECs, and addition of 17K did not result in any further decrease. Overexpression of uPAR in THP-1 macrophages leads to greater than a two fold increase in 17K and plasminogen binding. Plasminogen activation increases over two-fold as a result of overexpression of uPAR, while 17K blunts the effect of uPAR overexpression. These results indicate that uPAR plays a crucial role in both plasminogen and apo(a) binding to the cell surface of specific cells and inhibition by apo(a) of plasminogen activation. Macrophage-1-antigen (Mac-1) receptor consists of CD11b (αM) and CD18 (β2) integrin and has been previously shown to recognize uPA and control migration and adhesion. Furthermore, αVβ3 has been previously shown to bind to vitronectin and the uPA-uPAR complex which promotes cell adhesion through binding of both vitronectin and αVβ3 integrins. We found that blocking the αM, β2, or αVβ3 receptors with monoclonal antibodies in THP-1 cells leads to a decrease in plasminogen activation, as well as a blunting of the inhibitory effects of apo(a) on plasminogen activation. These results indicate a role for Mac-1 and αVβ3 in apo(a) binding and inhibition of plasminogen activation. In conclusion, we have demonstrated, for the first time, the role of specific receptors in binding of apo(a) to vascular cell surfaces and in mediating the inhibitory effect of apo(a) on pericellular plasminogen activation. Disclosures: No relevant conflicts of interest to declare.

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