Subcellular compartmentalization of saccharide moieties in cultured normal and malignant cells.

Virtanen, Ismo; Ekblom, Peter; Laurila, Pekka

doi:10.1083/jcb.85.2.429

Cited by 258 publications

(147 citation statements)

References 30 publications

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“…This lectin specifically binds to mannose-rich carbohydrate cores in glycoproteins and labels the ER [24,25]. Immunocytochemistry with concanavalin A revealed the net-like meshwork of the ER around the nucleus of the cells (Figure 4B), similar to the staining observed with the G594∆ mutant ( Figure 4A, lower panel).…”

Section: Immunocytochemical Localization Of the Apa Mutantssupporting

confidence: 53%

The C-terminal domain, but not the interchain disulphide, is required for the activity and intracellular trafficking of aminopeptidase A

Ofner¹,

Hooper²

2002

Biochem. J.

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Section: Immunocytochemical Localization Of the Apa Mutantssupporting

confidence: 53%

The C-terminal domain, but not the interchain disulphide, is required for the activity and intracellular trafficking of aminopeptidase A

Ofner¹,

Hooper²

2002

Biochem. J.

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“…However, we observed no differences in the efficiency of immunoprecipitation of the wildtype or glycosylation mutants when the affinity-purified anti-G protein antibody was used (data not shown). The internal staining pattern of the mutant G protein encoded by pTA1,2 obtained with the anti-G protein antibody was also compared with the pattern observed after the cells were stained with wheat germ agglutinin, which preferentially localizes in the Golgi complex due to its binding of terminal N-acetylglucosamine residues (47). The perinuclear regions stained by the anti-G protein antibody in cells transfected with pTA1,2 ( Fig.…”

Section: Resultsmentioning

confidence: 99%

A single N-linked oligosaccharide at either of the two normal sites is sufficient for transport of vesicular stomatitis virus G protein to the cell surface.

Machamer¹,

Florkiewicz²,

Rose³

1985

Mol. Cell. Biol.

159

114

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We investigated the role of glycosylation in intracellular transport and cell surface expression of the vesicular stomatitis virus glycoprotein (G) in cells expressing G protein from cloned cDNA. The individual contributions of the two asparagine-linked glycans of G protein to cell surface expression were assessed by site-directed mutagenesis of the coding sequence to eliminate one or the other or both of the glycosylation sites. One oligosaccharide at either position was sufficient for cell surface expression of G protein in transfected cells, and the rates of oligosaccharide processing were similar to the rate observed for wild-type protein. However, the nonglycosylated G protein synthesized when both glycosylation sites were eliminated did not reach the cell surface. This protein did appear to reach a Golgi-like region, as determined by indirect immunofluorescence microscopy, however, and was modified with palmitic acid. It was also apparently not subject to increased proteolytic breakdown.The carbohydrate moieties of glycoproteins have long been postulated to provide some function for the polypeptides to which they are covalently linked. Functions which have been implicated include maintenance of correct polypeptide conformation, protection from proteolytic degradation, and signals for intracellular targeting (reviewed in reference 29). Many experiments analyzing the role of asparagine-linked (N-linked) oligosaccharides on glycoproteins have used the antibiotic tunicamycin, which interferes with the synthesis of the lipid-linked precursor oligosaccharide and thus prevents addition of the core glycan to the nascent polypeptide (44, 46). The results of such experiments indicate that there is a wide range in the requirement for carbohydrates on secreted and integral membrane glycoproteins. Some proteins which normally contain N-linked oligosaccharides are not affected when they are synthesized in the presence of tunicamycin, whereas others fail to reach their correct cellular destination or are degraded (29).The glycoprotein (G protein) of vesicular stomatitis virus (VSV) has been extensively studied as a model for glycoprotein biosynthesis. This polypeptide is an integral membrane protein which contains 511 amino acids and the following three domains: a large amino-terminal external domain, a transmembrane domain of 20 amino acids, and a highly charged cytoplasmic domain of 29 amino acids (34). In infected cells, G protein is synthesized on membrane-bound ribosomes, and two high-mannose oligosaccharides are added cotranslationally as the polypeptide is extruded into the lumen of the rough endoplasmic reticulum (rER) (35). Translocation of the protein into the lumen stops at the hydrophobic transmembrane domain, and the membraneanchored polypeptide moves through the Golgi complex to the plasma membrane, where virus binding occurs (2,4,18). Several posttranslational modifications of G protein can be followed as the protein moves through the cell. These include the addition of palmitic acid to a cysteine residue in

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“…However, when the F-Con A staining is not too intense, one can notice that F-Con A is never present at this site in association with pp60 ~-'~ (thin white arrows, Fig. 9, e and f); (e) if the cells treated with F-Con A for 1 h are stained in parallel with fluorescent wheat germ agglutinin, which reveals the Golgi apparatus (Virtanen et al, 1980), one observes a coincidence between the F-Con A cap and that intracellular structure (arrows, Fig. 9, q and r).…”

Section: Simultaneous Observation Of the Pp60~ And Con A Patterns In mentioning

confidence: 99%

“…We next compared the intracellular distributions of pp60 ~-sr~ in the NIH (pM c-src/focus)B cells and of two labeled leetins, FITC-wheat germ agglutinin and TRITC-Con A, that preferentially label respectively the Golgi apparatus and the rough (R) ER (Virtanen et al, 1980). The three exhibit a perinuclear location, so it is not surprising that they appear to partially colocalize.…”

Section: Comparison Between the Localization Patterns Of Pp60 ~'~ Andmentioning

confidence: 99%

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Immunolocalization of the cellular src protein in interphase and mitotic NIH c-src overexpresser cells.

David‐Pfeuty

Nouvian-Dooghe

1990

The Journal of Cell Biology

129

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The mouse mAb, mAb 327, that recognizes specifically both pp60v-src and pp60c-src in a wide variety of cells, has been used to determine precisely the various locations of pp60c-src in NIH c-src overexpresser cells, using the technique of immunofluorescence microscopy. In interphase cells, the protein exhibits two main distributions: one that appears uniform and in association with the cell surface and the other that is patchy and juxtanuclear and coincides with the centrosomes. The juxtanuclear aggregation of pp60c-src-containing patches depends on microtubules and does not seem to occur within the Golgi apparatus and the rough ER. At the G2-to-M-phase transition, a drastic change in the localization patterns of pp60c-src takes place. We also report experiments in which the NIH c-src overexpresser cells were exposed to Con A for various times to induce a redistribution of the cell surface Con A receptors. We show that, at each stage of the Con A-mediated endocytotic process, the Con A-receptor complexes redistribute into structures to which pp60c-src appears also to be associated: at first, into patches that form at the cell surface level and then, into a cap that stands at the cell center in a juxtanuclear position and that coincides with the Golgi apparatus. During this capping process, pp60c-src-containing vesicles continue to accumulate in a centriolar spot, as in interphase, Con A-untreated cells, from which Con A is excluded. The significance of the intracellular locations of pp60c-src to the possible functions of the protein is discussed. Also, the distribution patterns of the cellular protein in the NIH c-src overexpresser cells are compared with those of pp60v-src in RSV-transformed cells. The differences observed are discussed in relation with the differences in transforming capacities of the two proteins. Finally, the possible physiological significance of the association between pp60c-src and the structures generated after the binding of Con A to its surface receptors is addressed.

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Subcellular compartmentalization of saccharide moieties in cultured normal and malignant cells.

Cited by 258 publications

References 30 publications

The C-terminal domain, but not the interchain disulphide, is required for the activity and intracellular trafficking of aminopeptidase A

The C-terminal domain, but not the interchain disulphide, is required for the activity and intracellular trafficking of aminopeptidase A

A single N-linked oligosaccharide at either of the two normal sites is sufficient for transport of vesicular stomatitis virus G protein to the cell surface.

Immunolocalization of the cellular src protein in interphase and mitotic NIH c-src overexpresser cells.

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