Molecular Characterisation and Structural Relationship of the Synapse‐Enriched Glycoproteins gp65 and gp55

Willmott, Tim; Skitsa, Ioulia; Hill, Irene E.; Mummery, Rosemary S.; Beesley, Philip

doi:10.1111/j.1471-4159.1992.tb10944.x

Cited by 18 publications

(32 citation statements)

References 24 publications

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“…The gp65-like immunoreactivity is specifically localized to the processes and nerve terminals of subsets of forebrain neurons. Biochemical studies suggest that the two molecules contain similar amounts of N-linked oligosaccharide and differ by a single extracellular 10 -12-kDa peptide sequence that contains little or no carbohydrate (12). The present data demonstrate that gp65 and gp55 are novel members of the Ig superfamily containing three and two Ig domains, respectively.…”

mentioning

confidence: 55%

Synaptic Membrane Glycoproteins gp65 and gp55 Are New Members of the Immunoglobulin Superfamily

Langnaese

Beesley

Gundelfinger

1997

Journal of Biological Chemistry

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104

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Glycoproteins gp65 and gp55 are major components of synaptic membranes prepared from rat forebrain. Both are recognized by the monoclonal antibody SMgp65. We have used SMgp65 to screen a rat brain cDNA expression library. Two sets of overlapping cDNAs that contain open reading frames of 397 and 281 amino acids were isolated. The deduced proteins are members of the immunoglobulin (Ig) superfamily containing three and two Ig domains, respectively. The common part has ϳ40% sequence identity with neurothelin/basigin. The identity of the proteins as gp65 and gp55 was confirmed by production of new antisera against a common recombinant protein fragment. These antisera immunoprecipitate gp65 and gp55. Furthermore, expression of gp65 and gp55 cDNAs in human 293 cells treated with tunicamycin results in the production of unglycosylated core proteins of identical size to deglycosylated gp65 and gp55. Northern analysis revealed that gp65 transcripts are brain-specific, whereas gp55 is expressed in most tissues and cell lines examined. The tissue distribution was confirmed at the protein level though the pattern of glycosylation of gp55 varies between tissues. In situ hybridization experiments with a common and a gp65-specific probe suggest differential expression of gp65 and gp55 transcripts in the rat brain.Synaptic junctions are highly specialized areas of contact and communication between neurons. They comprise the membranes and the underlying cytoskeleton of the pre-and postsynaptic neurons. In particular, the postsynaptic nerve ending is characterized by the presence of an electron-dense cytoskeletal structure, the postsynaptic density (PSD) 1 , which underlies the postsynaptic membrane. It is well established that synaptic structures and biochemically isolated synaptic subfractions, particularly synaptic membrane (SM) and PSD fractions, are enriched in distinct sets of glycoproteins that bind the lectin concanavalin A (1, 2). Some of these glycoproteins have been identified and their functions established. These include the major 180-kDa PSD glycoprotein that corresponds to the NR2B subunit of the N-methyl-D-aspartate receptor (3) and the SMenriched glycoprotein, gp50, that has been identified as the ␤2 subunit of the Na ϩ /K ϩ -ATPase (4, 5). SM and PSD glycoproteins, which are oriented with their oligosaccharide-containing domains facing into the synaptic cleft, are ideally placed to mediate adhesive interactions between the pre-and postsynaptic nerve endings. Indeed, the PSD-enriched PAC 1 glycoproteins (6, 7) have been identified as members of the cadherin family (8), providing evidence for such a role. Furthermore, integrin-type adhesion molecules (9) and NCAM (10), a member of the immunoglobulin (Ig) superfamily, have been recognized as SM components. Considering the complex interplay between stabilization and plasticity at synaptic connections, it is likely that more synaptic glycoproteins with adhesive functions are to be identified.Here we describe the molecular cloning of cDNAs encoding two synaptic glycopr...

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mentioning

confidence: 55%

Synaptic Membrane Glycoproteins gp65 and gp55 Are New Members of the Immunoglobulin Superfamily

Langnaese

Beesley

Gundelfinger

1997

Journal of Biological Chemistry

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104

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“…Further, Bsg shows homology with two other Ig superfamily members, embigin (28% homology) and gp65/gp55 (40±44% homology). The former was found in embryonal carcinoma cells [35], and exhibits a cell-substratum adhesion-enhancing activity [36], whereas the latter was found as synaptic membrane glycoproteins [37,38]. A striking structural feature of these three proteins is again that there is a charged residue, glutamic acid, at a similar position in the single membrane-spanning region [10,35,38].…”

Section: Discussionmentioning

confidence: 99%

Homo‐oligomer formation by basigin, an immunoglobulin superfamily member, via its N‐terminal immunoglobulin domain

Yoshida¹,

Shibata²,

Yamamoto³

et al. 2000

European Journal of Biochemistry

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Basigin (Bsg) is a highly glycosylated transmembrane protein with two immunoglobulin (Ig)-like domains. A number of studies, including gene targeting, have demonstrated that Bsg plays pivotal roles in spermatogenesis, implantation, neural network formation and tumor progression. In the present study, to understand the mechanism of action of Bsg, we determined its expression status on the plasma membrane. Cotransfection of Bsg expression vectors with two different tags clarified that Bsg forms homo-oligomers in a cis-dependent manner on the plasma membrane. If the disulfide bond of the more N-terminally located Ig-like domain was destroyed by mutations, Bsg could not form oligomers. In contrast, the mutations of the C-terminal Ig-like domain or N-glycosylation sites did not affect the association. The association of mouse and human Bsgs, which exhibit high homology in the transmembrane and intracellular domains but low homology in the extracellular domain, was very weak as compared with that within the same species, suggesting the importance of the extracellular domain in the association. If the extracellular domain of the human Ret protein was replaced with the N-terminal Ig-like domain of Bsg, the resulting chimera protein was associated with intact wild-type Bsg, but not if the C-terminal Ig-like domain, instead of the N-terminal one, of Bsg was used. No oligomer formation took place between the intact wild-type Ret and Bsg proteins. In conclusion, these data indicate that the N-terminal Ig-like domain is necessary and sufficient for oligomer formation by Bsg on the plasma membrane.Keywords: basigin; EMMPRIN; immunoglobulin superfamily; neurothelin; oligomer.Cell-surface proteins, such as the Ig superfamily, the cadherin superfamily and the integrin superfamily, play important roles in development by regulating cellular differentiation, adhesion and migration [1,2]. Supramolecular structure formation, such as oligomer formation, between these molecules plays crucial roles in exerting their functions. This has been documented by extensive studies of the Ig superfamily. NCAM participates in neural cell adhesion, which is mainly mediated by homophilic binding in a trans-dependent manner through its third Ig-like domain [3,4]. Homodimer formation of ICAM in a cis-dependent manner enhances its binding to a leukocyte b2-integrin [5]. Binding of fibroblast growth factor (FGF) to its receptor (FGFR) leads to dimer formation of FGFR (eventually a complex formation consisting of two FGF, two FGFR and a heparan sulfate chain), which is mediated mainly through homophilic binding at the second Ig-like domain of FGFR [6]. In addition to protein±protein interaction as mentioned above, protein±sugar interaction is also important. P0, a neural adhesion molecule implicated in myelination, harbors an N-glycosylation site, to which the major homophilic binding site is mapped [7]. NCAM and L1, another neural adhesion molecule, associate each other in a cis-dependent manner to reinforce L1 homophilic trans-interaction: in this case...

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“…Western blots were immunodeveloped by overnight incubation with primary antibody and processed employing the ECL detection system (Amersham Corp. Phospholipase treatment was carried out using 0.6 units of PI-PLC (Oxford GlycoSystems) and 0.03 units of chondroitinase ABC/30 g of membrane proteins as described by Ref. 29.…”

Section: Methodsmentioning

confidence: 99%

Brevican, a Chondroitin Sulfate Proteoglycan of Rat Brain, Occurs as Secreted and Cell Surface Glycosylphosphatidylinositol-anchored Isoforms

Seidenbecher

Richter

Rauch

et al. 1995

Journal of Biological Chemistry

141

134

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cDNA clones encoding proteins related to the aggrecan/versican family of proteoglycan core proteins have been isolated with antisera against rat brain synaptic junctions. Two sets of overlapping cDNAs have been characterized that differ in their 3-terminal regions. Northern analyses with probes derived from unique regions of each set were found to hybridize with two brain-specific transcripts of 3.3 and 3.6 kilobases (kb). The 3.6-kb transcript encodes a polypeptide that exhibits 82% sequence identity with bovine brevican and is thought to be the rat ortholog of brevican. Interestingly, the polypeptide deduced from the open reading frame of the 3.3-kb transcript is truncated just carboxyl-terminal of the central domain of brevican and instead contains a putative glypiation signal. Antibodies raised against a bacterially expressed glutathione S-transferase-brevican fusion protein have been used to show that both soluble and membrane-bound brevican isoforms exist. Treatment of the crude membrane fraction and purified synaptic plasma membranes with phosphatidylinositolspecific phospholipase C revealed that isoforms of brevican are indeed glycosylphosphatidylinositol-anchored to the plasma membrane. Moreover, digestions with chondroitinase ABC have indicated that rat brevican, like its bovine ortholog, is a conditional chondroitin sulfate proteoglycan. Immunohistochemical studies have shown that brevican is widely distributed in the brain and is localized extracellularly. During postnatal development, amounts of both soluble and phosphatidylinositol-specific phospholipase C-sensitive isoforms increase, suggesting a role for brevican in the terminally differentiating and the adult nervous system.

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Molecular Characterisation and Structural Relationship of the Synapse‐Enriched Glycoproteins gp65 and gp55

Cited by 18 publications

References 24 publications

Synaptic Membrane Glycoproteins gp65 and gp55 Are New Members of the Immunoglobulin Superfamily

Synaptic Membrane Glycoproteins gp65 and gp55 Are New Members of the Immunoglobulin Superfamily

Homo‐oligomer formation by basigin, an immunoglobulin superfamily member, via its N‐terminal immunoglobulin domain

Brevican, a Chondroitin Sulfate Proteoglycan of Rat Brain, Occurs as Secreted and Cell Surface Glycosylphosphatidylinositol-anchored Isoforms

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