Characterization of a Cell Surface Protein ofClostridium difficile with Adhesive Properties

ABSTRACT Our laboratory has previously shown that Clostridium difficile adherence to cultured cells is enhanced after heat shock at 60°C and that it is mediated by a proteinaceous surface component. The present study was undertaken to identify the surface molecules of this bacterium that could play a role in its adherence to the intestine. The cwp66 gene, encoding a cell surface-associated protein of C. difficile 79-685, was isolated by immunoscreening of a C. difficile gene library with polyclonal antibodies against C. difficile heated at 60°C. The Cwp66 protein (66 kDa) contains two domains, each carrying three imperfect repeats and one presenting homologies to the autolysin CwlB of Bacillus subtilis. A survey of 36 strains ofC. difficile representing 11 serogroups showed that the 3′ portion of the cwp66 gene is variable; this was confirmed by sequencing of cwp66 from another strain, C-253. Two recombinant protein fragments corresponding to the two domains of Cwp66 were expressed in fusion with glutathione S-transferase inEscherichia coli and purified by affinity chromatography using gluthatione-Sepharose 4B. Antibodies raised against the two domains recognized Cwp66 in bacterial surface extracts. By immunoelectron microscopy, the C-terminal domain was found to be cell surface exposed. When used as inhibitors in cell binding studies, the antibodies and protein fragments partially inhibited adherence ofC. difficile to cultured cells, confirming that Cwp66 is an adhesin, the first to be identified in clostridia.

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Functional correlation between cell adhesive properties and some cell surface proteins.

The Journal of Cell Biology ◽

10.1083/jcb.75.2.464 ◽

1977 ◽

Vol 75 (2) ◽

pp. 464-474 ◽

Cited By ~ 390

Author(s):

M Takeichi

Keyword(s):

Cell Surface ◽

Chinese Hamster ◽

Surface Protein ◽

Surface Proteins ◽

Adhesive Properties ◽

Cell Surface Proteins ◽

Morphological Studies ◽

Chinese Hamster V79 Cells ◽

A Cell ◽

In Cells

The adhesive properties of Chinese hamster V79 cells were analyzed and characterized by various cell dissociation treatments. The comparisons of aggregability among cells dissociated with EDTA, trypsin + Ca2+, and trypsin + EDTA, revealed that these cells have two adhesion mechanisms, a Ca2+-independent and a Ca2+-dependent one. The former did not depend on temperature, whereas the latter occurred only at physiological temperatures. Both mechanisms were trypsin sensitive, but the Ca2+-dependent one was protected by Ca2+ against trypsinization. In morphological studies, the Ca2+-independent adhesion appeared to be a simple agglutination or flocculation of cells, whereas the Ca2+-dependent adhesion seemed to be more physiological, being accompanied by cell deformation resulting in the increase of contact area between adjacent cells. Lactoperoxidase-catalyzed iodination of cell surface proteins revealed that several proteins are more intensely labeled in cells with Ca2+-independent adhesiveness than in cells without that property. It was also found that a cell surface protein with a molecular weight of approximately 150,000 is present only in cells with Ca2+-dependent adhesiveness. The iodination and trypsinization of this protein were protected by Ca2+, suggesting its reactivity to Ca2+. Possible mechanisms for each adhesion property are discussed, taking into account the correlation of these proteins with cell adhesiveness.

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Identification of PA2.26 antigen as a novel cell-surface mucin-type glycoprotein that induces plasma membrane extensions and increased motility in keratinocytes

Journal of Cell Science ◽

10.1242/jcs.112.24.4601 ◽

1999 ◽

Vol 112 (24) ◽

pp. 4601-4613 ◽

Cited By ~ 1

Author(s):

F.G. Scholl ◽

C. Gamallo ◽

S. Vilar?o ◽

M. Quintanilla

Keyword(s):

Plasma Membrane ◽

Cell Surface ◽

Biochemical Characterization ◽

Cultured Cells ◽

Ectopic Expression ◽

Lymphatic Vessels ◽

Surface Protein ◽

Cultured Fibroblasts ◽

Transmembrane Glycoprotein ◽

A Cell

PA2.26 antigen was identified as a cell-surface protein induced in epidermal carcinogenesis and skin remodeling processes. PA2.26 is expressed in carcinoma cell lines and cultured fibroblasts but absent in nontumorigenic keratinocytes. In tissues, PA2.26 is present in epithelial cells of the choroid plexus, ependyma, glomerulus and alveolus, in mesothelial cells, and in endothelia of lymphatic vessels. Biochemical characterization of PA2.26 protein and sequence analysis of the isolated cDNA demonstrate that PA2.26 antigen is a mucin-like transmembrane glycoprotein. Confocal and immunoelectron microscopy analysis in cultured cells reveal that PA2. 26 is concentrated in actin-rich microvilli and plasma membrane projections, such as filopodia, lamellipodia and ruffles, where it colocalizes with members of the ERM (ezrin, radixin, moesin) family protein. Ezrin and moesin, but not radixin, can be coimmunoprecipitated together with PA2.26 from cell lysates. Ectopic expression of PA2.26 in immortalized, nontumorigenic, keratinocytes induces an epithelial-fibroblastoid morphological conversion with increased plasma membrane extensions, concomitantly to a major reorganization of the actin cytoskeleton, redistribution of ezrin to cell-surface projections, and enhanced motility. These findings suggest an involvement of PA2.26 in cell migration.

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Leukocytes with chromosome Y loss have reduced abundance of the cell surface immunoprotein CD99

Scientific Reports ◽

10.1038/s41598-021-94588-5 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Jonas Mattisson ◽

Marcus Danielsson ◽

Maria Hammond ◽

Hanna Davies ◽

Caroline J. Gallant ◽

...

Keyword(s):

Cell Surface ◽

Single Cells ◽

Surface Protein ◽

Protein Abundance ◽

Blood Leukocytes ◽

Chromosome Y ◽

Increased Risk ◽

A Cell ◽

Pseudoautosomal Regions ◽

Male Specific

AbstractMosaic loss of chromosome Y (LOY) in immune cells is a male-specific mutation associated with increased risk for morbidity and mortality. The CD99 gene, positioned in the pseudoautosomal regions of chromosomes X and Y, encodes a cell surface protein essential for several key properties of leukocytes and immune system functions. Here we used CITE-seq for simultaneous quantification of CD99 derived mRNA and cell surface CD99 protein abundance in relation to LOY in single cells. The abundance of CD99 molecules was lower on the surfaces of LOY cells compared with cells without this aneuploidy in all six types of leukocytes studied, while the abundance of CD proteins encoded by genes located on autosomal chromosomes were independent from LOY. These results connect LOY in single cells with immune related cellular properties at the protein level, providing mechanistic insight regarding disease vulnerability in men affected with mosaic chromosome Y loss in blood leukocytes.

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Localization and expression of msp130, a primary mesenchyme lineage-specific cell surface protein in the sea urchin embryo

Development ◽

10.1242/dev.101.2.255 ◽

1987 ◽

Vol 101 (2) ◽

pp. 255-265 ◽

Cited By ~ 4

Author(s):

J.A. Anstrom ◽

J.E. Chin ◽

D.S. Leaf ◽

A.L. Parks ◽

R.A. Raff

Keyword(s):

Cell Surface ◽

Sea Urchin ◽

Sea Water ◽

Surface Protein ◽

Specific Cell ◽

Cell Surface Protein ◽

Sea Urchin Embryo ◽

A Cell ◽

Mesenchyme Cells ◽

Primary Mesenchyme Cells

In this report, we use a monoclonal antibody (B2C2) and antibodies against a fusion protein (Leaf et al. 1987) to characterize msp130, a cell surface protein specific to the primary mesenchyme cells of the sea urchin embryo. This protein first appears on the surface of these cells upon ingression into the blastocoel. Immunoelectronmicroscopy shows that msp130 is present in the trans side of the Golgi apparatus and on the extracellular surface of primary mesenchyme cells. Four precursor proteins to msp130 are identified and we show that B2C2 recognizes only the mature form of msp130. We demonstrate that msp130 contains N-linked carbohydrate groups and that the B2C2 epitope is sensitive to endoglycosidase F digestion. Evidence that msp130 is apparently a sulphated glycoprotein is presented. The recognition of the B2C2 epitope of msp130 is disrupted when embryos are cultured in sulphate-free sea water. In addition, two-dimensional immunoblots show that msp130 is an acidic protein that becomes substantially less acidic in the absence of sulphate. We also show that two other independently derived monoclonal antibodies, IG8 (McClay et al. 1983; McClay, Matranga & Wessel, 1985) and 1223 (Carson et al. 1985), recognize msp130, and suggest this protein to be a major cell surface antigen of primary mesenchyme cells.

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A cell surface protein controls endocrine ring gland morphogenesis and steroid production

Developmental Biology ◽

10.1016/j.ydbio.2018.10.007 ◽

2019 ◽

Vol 445 (1) ◽

pp. 16-28 ◽

Cited By ~ 2

Author(s):

Yanina-Yasmin Pesch ◽

Ricarda Hesse ◽

Tariq Ali ◽

Matthias Behr

Keyword(s):

Cell Surface ◽

Surface Protein ◽

Cell Surface Protein ◽

Steroid Production ◽

Ring Gland ◽

A Cell ◽

Gland Morphogenesis

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The in silico human surfaceome

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1808790115 ◽

2018 ◽

Vol 115 (46) ◽

pp. E10988-E10997 ◽

Cited By ~ 44

Author(s):

Damaris Bausch-Fluck ◽

Ulrich Goldmann ◽

Sebastian Müller ◽

Marc van Oostrum ◽

Maik Müller ◽

...

Keyword(s):

Cell Surface ◽

In Silico ◽

Embryonic Stem ◽

Surface Protein ◽

Surface Proteins ◽

Cell Surface Protein ◽

Cell Surface Proteins ◽

A Cell ◽

Visualization Tools ◽

Protein Classes

Cell-surface proteins are of great biomedical importance, as demonstrated by the fact that 66% of approved human drugs listed in the DrugBank database target a cell-surface protein. Despite this biomedical relevance, there has been no comprehensive assessment of the human surfaceome, and only a fraction of the predicted 5,000 human transmembrane proteins have been shown to be located at the plasma membrane. To enable analysis of the human surfaceome, we developed the surfaceome predictor SURFY, based on machine learning. As a training set, we used experimentally verified high-confidence cell-surface proteins from the Cell Surface Protein Atlas (CSPA) and trained a random forest classifier on 131 features per protein and, specifically, per topological domain. SURFY was used to predict a human surfaceome of 2,886 proteins with an accuracy of 93.5%, which shows excellent overlap with known cell-surface protein classes (i.e., receptors). In deposited mRNA data, we found that between 543 and 1,100 surfaceome genes were expressed in cancer cell lines and maximally 1,700 surfaceome genes were expressed in embryonic stem cells and derivative lines. Thus, the surfaceome diversity depends on cell type and appears to be more dynamic than the nonsurface proteome. To make the predicted surfaceome readily accessible to the research community, we provide visualization tools for intuitive interrogation (wlab.ethz.ch/surfaceome). The in silico surfaceome enables the filtering of data generated by multiomics screens and supports the elucidation of the surfaceome nanoscale organization.

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