The dual role of amyloid-β-sheet sequences in the cell surface properties of FLO11-encoded flocculins in Saccharomyces cerevisiae

Fungal adhesins (Als) or flocculins are family of cell surface proteins that mediate adhesion to diverse biotic and abiotic surfaces. A striking characteristic of Als proteins originally identified in the pathogenic Candida albicans is to form functional amyloids that mediate cis-interaction leading to the formation of adhesin nanodomains and trans-interaction between amyloid sequences of opposing cells. In this report, we show that flocculins encoded by FLO11 in Saccharomyces cerevisiae behave like adhesins in C. albicans. To do so, we show that the formation of nanodomains under an external physical force requires a threshold number of amyloid-forming sequences in the Flo11 protein. Then, using a genome editing approach, we constructed strains expressing variants of the Flo11 protein under the endogenous FLO11 promoter, leading to the demonstration that the loss of amyloid-forming sequences strongly reduces cell-cell interaction but has no effect on either plastic adherence or invasive growth in agar, both phenotypes being dependent on the N- and C-terminal ends of Flo11p. Finally, we show that the location of Flo11 is not altered either by the absence of amyloid-forming sequences or by the removal of the N- or C-terminus of the protein.

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Role of cations in the flocculation of Saccharomyces cerevisiae and discrimination of the corresponding proteins

Canadian Journal of Microbiology ◽

10.1139/m91-064 ◽

1991 ◽

Vol 37 (5) ◽

pp. 397-403 ◽

Cited By ~ 17

Author(s):

Hiroshi Kuriyama ◽

Itaru Umeda ◽

Harumi Kobayashi

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Surface ◽

Inhibitory Effect ◽

Surface Proteins ◽

Promoting Effect ◽

Yeast Strains ◽

Yeast Flocculation ◽

Different Types ◽

Anionic Groups

Asexual yeast flocculation was studied using strong flocculents of Saccharomyces cerevisiae. The inhibitory effect of cations on flocculation is considered to be caused by competition between those cations and Ca2+ at the binding site of the Ca2+-requiring protein that is involved in flocculation. Inhibition of flocculation by various cations occurred in the following order: La3+, Sr2+, Ba2+, Mn2+, Al3+, and Na+. Cations such as Mg2+, Co2+, and K+ promoted flocculation. This promoting effect may be based on the reduction of electrostatic repulsive force between cells caused by binding of these cations anionic groups present on the cell surface. In flocculation induced by these cations, trace amounts of Ca2+ excreted on the cell surface may activate the corresponding protein. The ratio of Sr2+/Ca2+ below which cells flocculated varied among strains: for strains having the FLO5 gene, it was 400 to 500; for strains having the FLO1 gene, about 150; and for two alcohol yeast strains, 40 to 50. This suggests that there are several different types of cell surface proteins involved in flocculation in different yeast strains. Key words: yeast, flocculation, protein, cation, calcium.

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Gel and gel-free proteomics to identify Saccharomyces cerevisiae cell surface proteins

Journal of Proteomics ◽

10.1016/j.jprot.2010.02.005 ◽

2010 ◽

Vol 73 (6) ◽

pp. 1183-1195 ◽

Cited By ~ 35

Author(s):

María Rosa Insenser ◽

María Luisa Hernáez ◽

César Nombela ◽

María Molina ◽

Gloria Molero ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Surface ◽

Surface Proteins ◽

Cell Surface Proteins

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Intracellular nanovesicles mediate integrin trafficking during cell migration

10.1101/2020.08.19.257287 ◽

2020 ◽

Author(s):

Gabrielle Larocque ◽

Penelope J. La-Borde ◽

Beverley J. Wilson ◽

Nicholas I. Clarke ◽

Daniel J. Moore ◽

...

Keyword(s):

Cell Migration ◽

Cell Surface ◽

Membrane Trafficking ◽

Surface Proteins ◽

Migration And Invasion ◽

Rab Gtpase ◽

Surface Receptors ◽

C Terminus ◽

Transport Vesicle ◽

Important Regulator

Membrane traffic is an important regulator of cell migration through the endocytosis and recycling of cell surface receptors such as integrin heterodimers. Intracellular nanovesicles (INVs), are a recently identified class of transport vesicle that are involved in multiple membrane trafficking steps including the recycling pathway. The only known marker for INVs is Tumor Protein D54 (TPD54/TPD52L2), a member of the TPD52-like protein family. Overexpression of TPD52-like family proteins in cancer has been linked to poor prognosis and an aggressive metastatic phenotype which suggests cell migration may be altered under these conditions. Here we show that TPD54 associates with INVs by directly binding high curvature membrane via a conserved positively charged motif in its C-terminus. We describe how other members of the TPD52-like family are also associated with INVs and we document the Rab GTPase complement of all INVs. Depletion of TPD52-like proteins inhibits cell migration and invasion; and we show that this is likely due to altered integrin recycling. Our study highlights the involvement of INVs in the trafficking of cell surface proteins to generate biologically important outputs in health and disease.

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ArtA-Dependent Processing of a Tat Substrate Containing a Conserved Tripartite Structure That Is Not Localized at the C Terminus

Journal of Bacteriology ◽

10.1128/jb.00802-16 ◽

2017 ◽

Vol 199 (7) ◽

Cited By ~ 7

Author(s):

Mohd Farid Abdul Halim ◽

Jonathan D. Stoltzfus ◽

Stefan Schulze ◽

Micheal Hippler ◽

Mechthild Pohlschroder

Keyword(s):

Cell Surface ◽

In Silico ◽

Cell Biology ◽

Surface Proteins ◽

Signal Peptidase ◽

Secreted Proteins ◽

Dependent Manner ◽

Content Type ◽

C Terminus ◽

Signal Peptidase I

ABSTRACT Most prokaryote-secreted proteins are transported to the cell surface using either the general secretion (Sec) or twin-arginine translocation (Tat) pathway. A majority of secreted proteins are anchored to the cell surface, while the remainder are released into the extracellular environment. The anchored surface proteins play a variety of important roles in cellular processes, ranging from facilitating interactions between cells to maintaining cell stability. The extensively studied S-layer glycoprotein (SLG) of Haloferax volcanii, previously thought to be anchored via C-terminal intercalation into the membrane, was recently shown to be lipidated and to have its C-terminal segment removed in processes dependent upon archaeosortase A (ArtA), a recently discovered enzyme. While SLG is a Sec substrate, in silico analyses presented here reveal that, of eight additional ArtA substrates predicted, two substrates also contain predicted Tat signal peptides, including Hvo_0405, which has a highly conserved tripartite structure that lies closer to the center of the protein than to its C terminus, unlike other predicted ArtA substrates identified to date. We demonstrate that, even given its atypical location, this tripartite structure, which likely resulted from the fusion of genes encoding an ArtA substrate and a cytoplasmic protein, is processed in an ArtA-dependent manner. Using an Hvo_0405 mutant lacking the conserved “twin” arginines of the predicted Tat signal peptide, we show that Hvo_0405 is indeed a Tat substrate and that ArtA substrates include both Sec and Tat substrates. Finally, we confirmed the Tat-dependent localization and signal peptidase I (SPase I) cleavage site of Hvo_0405 using mass spectrometry. IMPORTANCE The specific mechanisms that facilitate protein anchoring to the archaeal cell surface remain poorly understood. Here, we have shown that the proteins bound to the cell surface of the model archaeon H. volcanii, through a recently discovered novel ArtA-dependent anchoring mechanism, are more structurally diverse than was previously known. Specifically, our results demonstrate that both Tat and Sec substrates, which contain the conserved tripartite structure of predicted ArtA substrates, can be processed in an ArtA-dependent manner and that the tripartite structure need not lie near the C terminus for this processing to occur. These data improve our understanding of archaeal cell biology and are invaluable for in silico subcellular localization predictions of archaeal and bacterial proteins.

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Gis4, a New Component of the Ion Homeostasis System in the Yeast Saccharomyces cerevisiae

Eukaryotic Cell ◽

10.1128/ec.00215-06 ◽

2006 ◽

Vol 5 (10) ◽

pp. 1611-1621 ◽

Cited By ~ 14

Author(s):

Tian Ye ◽

Raúl García-Salcedo ◽

José Ramos ◽

Stefan Hohmann

Keyword(s):

Saccharomyces Cerevisiae ◽

Salt Tolerance ◽

Cell Surface ◽

Ion Homeostasis ◽

Yeast Saccharomyces Cerevisiae ◽

Snf1 Kinase ◽

C Terminus ◽

Surface Localization ◽

As Stress

ABSTRACT Gis4 is a new component of the system required for acquisition of salt tolerance in Saccharomyces cerevisiae. The gis4Δ mutant is sensitive to Na+ and Li+ ions but not to osmotic stress. Genetic evidence suggests that Gis4 mediates its function in salt tolerance, at least partly, together with the Snf1 protein kinase and in parallel with the calcineurin protein phosphatase. When exposed to salt stress, mutants lacking gis4Δ display a defect in maintaining low intracellular levels of Na+ and Li+ ions and exporting those ions from the cell. This defect is due to diminished expression of the ENA1 gene, which encodes the Na+ and Li+ export pump. The protein sequence of Gis4 is poorly conserved and does not reveal any hints to its molecular function. Gis4 is enriched at the cell surface, probably due to C-terminal farnesylation. The CAAX box at the C terminus is required for cell surface localization but does not seem to be strictly essential for the function of Gis4 in salt tolerance. Gis4 and Snf1 seem to share functions in the control of ion homeostasis and ENA1 expression but not in glucose derepression, the best known role of Snf1. Together with additional evidence that links Gis4 genetically and physically to Snf1, it appears that Gis4 may function in a pathway in which Snf1 plays a specific role in controlling ion homeostasis. Hence, it appears that the conserved Snf1 kinase plays roles in different pathways controlling nutrient as well as stress response.

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Discovery of Fibrillar Adhesins across Bacterial Species

10.1101/2020.12.07.414375 ◽

2020 ◽

Author(s):

Vivian Monzon ◽

Aleix Lafita ◽

Alex Bateman

Keyword(s):

Cell Surface ◽

Bacterial Species ◽

Domain Architecture ◽

Surface Proteins ◽

Host Cells ◽

Bacterial Surface ◽

Bacterial Phyla ◽

C Terminus ◽

Wide Range ◽

Bacterial Proteomes

AbstractBackgroundFibrillar adhesins are long multidomain proteins attached at the cell surface and composed of at least one adhesive domain and multiple tandemly repeated domains, which build an elongated stalk that projects the adhesive domain beyond the bacterial cell surface. They are an important yet understudied class of proteins that mediate interactions of bacteria with their environment. This study aims to characterize fibrillar adhesins in a wide range of bacterial phyla and to identify new fibrillar adhesin-like proteins to improve our understanding of host-bacteria interactions.ResultsBy careful search for fibrillar adhesins in the literature and by computational analysis we identified 75 stalk domains and 24 adhesive domains. Based on the presence of these domains in the UniProt Reference Proteomes database, we identified and analysed 3,388 fibrillar adhesin-like proteins across species of the most common bacterial phyla. We found that the bacterial proteomes with the highest fraction of fibrillar adhesins include several known pathogens. We further enumerate the adhesive and stalk domain combinations found in nature and demonstrate that fibrillar adhesins have complex and variable domain architectures, which differ across species. By analysing the domain architecture of fibrillar adhesins we show that in Gram positive bacteria adhesive domains are mostly positioned at the N-terminus of the protein with the cell surface anchor at the C-terminus, while their positions are more variable in Gram negative bacteria. We provide an open repository of fibrillar adhesin-like proteins and domains to facilitate downstream studies of this class of bacterial surface proteins.ConclusionThis study provides a domain-based characterization of fibrillar adhesins and demonstrates that they are widely found across the main bacterial phyla. We have discovered numerous novel fibrillar adhesins and improved the understanding of how pathogens might adhere to and subsequently invade into host cells.

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Biosynthesis of Galactofuranose in Kinetoplastids: Novel Therapeutic Targets for Treating Leishmaniasis and Chagas' Disease

Enzyme Research ◽

10.4061/2011/415976 ◽

2011 ◽

Vol 2011 ◽

pp. 1-13 ◽

Cited By ~ 26

Author(s):

Michelle Oppenheimer ◽

Ana L. Valenciano ◽

Pablo Sobrado

Keyword(s):

Cell Surface ◽

Drug Target ◽

Biosynthetic Pathway ◽

Cell Interaction ◽

Surface Proteins ◽

Chemical Mechanism ◽

Protozoan Parasites ◽

Homologous Enzyme ◽

Unique Chemical

Cell surface proteins of parasites play a role in pathogenesis by modulating mammalian cell recognition and cell adhesion during infection. β-Galactofuranose (Galf) is an important component of glycoproteins and glycolipids found on the cell surface of Leishmania spp. and Trypanosoma cruzi. β-Galf-containing glycans have been shown to be important in parasite-cell interaction and protection against oxidative stress. Here, we discuss the role of β-Galf in pathogenesis and recent studies on the Galf-biosynthetic enzymes: UDP-galactose 4′ epimerase (GalE), UDP-galactopyranose mutase (UGM), and UDP-galactofuranosyl transferase (GalfT). The central role in Galf formation, its unique chemical mechanism, and the absence of a homologous enzyme in humans identify UGM as the most attractive drug target in the β-Galf-biosynthetic pathway in protozoan parasites.

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Discovery of fibrillar adhesins across bacterial species

BMC Genomics ◽

10.1186/s12864-021-07586-2 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Vivian Monzon ◽

Aleix Lafita ◽

Alex Bateman

Keyword(s):

Cell Surface ◽

Bacterial Species ◽

Domain Architecture ◽

Surface Proteins ◽

Bacterial Surface ◽

Bacterial Phyla ◽

Gram Positive Bacteria ◽

Invasion Mechanisms ◽

C Terminus ◽

Wide Range

Abstract Background Fibrillar adhesins are long multidomain proteins that form filamentous structures at the cell surface of bacteria. They are an important yet understudied class of proteins composed of adhesive and stalk domains that mediate interactions of bacteria with their environment. This study aims to characterize fibrillar adhesins in a wide range of bacterial phyla and to identify new fibrillar adhesin-like proteins to improve our understanding of host-bacteria interactions. Results Through careful literature and computational searches, we identified 82 stalk and 27 adhesive domain families in fibrillar adhesins. Based on the presence of these domains in the UniProt Reference Proteomes database, we identified and analysed 3,542 fibrillar adhesin-like proteins across species of the most common bacterial phyla. We further enumerate the adhesive and stalk domain combinations found in nature and demonstrate that fibrillar adhesins have complex and variable domain architectures, which differ across species. By analysing the domain architecture of fibrillar adhesins, we show that in Gram positive bacteria, adhesive domains are mostly positioned at the N-terminus and cell surface anchors at the C-terminus of the protein, while their positions are more variable in Gram negative bacteria. We provide an open repository of fibrillar adhesin-like proteins and domains to enable further studies of this class of bacterial surface proteins. Conclusion This study provides a domain-based characterization of fibrillar adhesins and demonstrates that they are widely found in species across the main bacterial phyla. We have discovered numerous novel fibrillar adhesins and improved our understanding of pathogenic adhesion and invasion mechanisms.

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Post-translational processing targets functionally diverse proteins in Mycoplasma hyopneumoniae

Open Biology ◽

10.1098/rsob.150210 ◽

2016 ◽

Vol 6 (2) ◽

pp. 150210 ◽

Cited By ~ 33

Author(s):

Jessica L. Tacchi ◽

Benjamin B. A. Raymond ◽

Paul A. Haynes ◽

Iain J. Berry ◽

Michael Widjaja ◽

...

Keyword(s):

Cell Surface ◽

Bacterial Pathogen ◽

Surface Protein ◽

Mycoplasma Hyopneumoniae ◽

Surface Proteins ◽

Post Translational Modification ◽

A Genome ◽

Associated Proteins ◽

Epithelial Cell Surface ◽

Functionally Diverse

Mycoplasma hyopneumoniae is a genome-reduced, cell wall-less, bacterial pathogen with a predicted coding capacity of less than 700 proteins and is one of the smallest self-replicating pathogens. The cell surface of M. hyopneumoniae is extensively modified by processing events that target the P97 and P102 adhesin families. Here, we present analyses of the proteome of M. hyopneumoniae- type strain J using protein-centric approaches (one- and two-dimensional GeLC–MS/MS) that enabled us to focus on global processing events in this species. While these approaches only identified 52% of the predicted proteome (347 proteins), our analyses identified 35 surface-associated proteins with widely divergent functions that were targets of unusual endoproteolytic processing events, including cell adhesins, lipoproteins and proteins with canonical functions in the cytosol that moonlight on the cell surface. Affinity chromatography assays that separately used heparin, fibronectin, actin and host epithelial cell surface proteins as bait recovered cleavage products derived from these processed proteins, suggesting these fragments interact directly with the bait proteins and display previously unrecognized adhesive functions. We hypothesize that protein processing is underestimated as a post-translational modification in genome-reduced bacteria and prokaryotes more broadly, and represents an important mechanism for creating cell surface protein diversity.

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Metabolic precision labeling enables selective probing of O-linked N-acetylgalactosamine glycosylation

10.1101/2020.04.23.057208 ◽

2020 ◽

Cited By ~ 1

Author(s):

Marjoke F. Debets ◽

Omur Y. Tastan ◽

Simon P. Wisnovsky ◽

Stacy A. Malaker ◽

Nikolaos Angelis ◽

...

Keyword(s):

Cell Surface ◽

Secretory Pathway ◽

Protein Glycosylation ◽

Surface Proteins ◽

Major Type ◽

Knock Out ◽

Biological Assays ◽

Superresolution Microscopy ◽

A Genome ◽

Health And Disease

AbstractProtein glycosylation events that happen early in the secretory pathway are often dysregulated during tumorigenesis. These events can be probed, in principle, by monosaccharides with bioorthogonal tags that would ideally be specific for distinct glycan subtypes. However, metabolic interconversion into other monosaccharides drastically reduces such specificity in the living cell. Here, we use a structure-based design process to develop the monosaccharide probe GalNAzMe that is specific for cancer-relevant Ser/Thr-N-acetylgalactosamine (O-GalNAc) glycosylation. By virtue of a branched N-acylamide side chain, GalNAzMe is not interconverted by epimerization to the corresponding N-acetylglucosamine analog like conventional GalNAc-based probes. GalNAzMe enters O-GalNAc glycosylation but does not enter other major cell surface glycan types including Asn (N)-linked glycans. We equip cells with the capacity to biosynthesize the nucleotide-sugar donor UDP-GalNAzMe from a caged precursor. Tagged with a bioorthogonal azide group, GalNAzMe serves as an O-glycan specific reporter in superresolution microscopy, chemical glycoproteomics, a genome-wide CRISPR knock-out (KO) screen, and imaging of intestinal organoids. GalNAzMe is a precision tool that allows a detailed view into the biology of a major type of cancer-relevant protein glycosylation.Significance statementA large portion of all secreted and cell surface proteins in humans are modified by Ser/Thr(O)-linked glycosylation with N-acetylgalactosamine (GalNAc). While of fundamental importance in health and disease, O-GalNAc glycosylation is technically challenging to study because of a lack of specific tools to be used in biological assays. Here, we design an O-GalNAc specific reporter molecule termed GalNAzMe to selectively label O-GalNAc glycoproteins in living human cells. GalNAzMe is compatible with a range of experiments in quantitative biology to broaden our understanding of glycosylation. We further demonstrate that labeling is genetically programmable by expression of a mutant glycosyltransferase, allowing application even to experiments with low inherent sensitivity.

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