Fucosylation and protein glycosylation create functional receptors for cholera toxin

Wands, Amberlyn M.; Fujita, Akiko; McCombs, Janet E.; Cervin, Jakob; Dedic, Benjamin; Rodríguez, A. Carlos Blesa; Nischan, Nicole; Bond, Michelle; Mettlen, Marcel; Trudgian, David C.; Lemoff, Andrew; Quiding‐Järbrink, Marianne; Gustavsson, Bengt; Steentoft, Catharina; Clausen, Henrik; Mirzaei, Hamid; Teneberg, Susann; Yrlid, Ulf; Kohler, Jennifer J.

doi:10.7554/elife.09545

Cited by 88 publications

(126 citation statements)

References 72 publications

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“…[44] In addition, GM 1 is of very low abundance (0.0015–0.003 mol% of glycosphingolipids) in human small intestinal epithelial cells[45]; thus, a recent publication raised a question, whether GM 1 is sufficient to induce cholera toxin attachment. [46] In the reduction of dimensionality model, high-affinity receptors can serve as initiators, and then activate weak receptors, leading to higher retention of CTB on the cell surface. Thus, the overall CTB binding is not simply controlled by a single GM 1 receptor; the weakly binding receptors can contribute to CTB binding via reduction of dimensionality.…”

Section: Discussionmentioning

confidence: 99%

Hetero-multivalent binding of cholera toxin subunit B with glycolipid mixtures

Krishnan

Singla

Lee

et al. 2017

Colloids and Surfaces B: Biointerfaces

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GM1 has generally been considered as the major receptor that binds to cholera toxin subunit B (CTB) due to its low dissociation constant. However, using a unique nanocube sensor technology, we have shown that CTB can also bind to other glycolipid receptors, fucosyl-GM1 and GD1b. Additionally, we have demonstrated that GM2 can contribute to CTB binding if present in a glycolipid mixture with a strongly binding receptor (GM1/fucosyl-GM1/GD1b). This hetero-multivalent binding result was unintuitive because the interaction between CTB and pure GM2 is negligible. We hypothesized that the reduced dimensionality of CTB-GM2 binding events is a major cause of the observed CTB binding enhancement. Once CTB has attached to a strong receptor, subsequent binding events are confined to a 2D membrane surface. Therefore, even a weak GM2 receptor could now participate in second or higher binding events because its surface reaction rate can be up to 104 times higher than the bulk reaction rate. To test this hypothesis, we altered the surface reaction rate by modulating the fluidity and heterogeneity of the model membrane. Decreasing membrane fluidity reduced the binding cooperativity between GM2 and a strong receptor. Our findings indicated a new protein-receptor binding assay, that can mimic complex cell membrane environment more accurately, is required to explore the inherent hetero-multivalency of the cell membrane. We have thus developed a new membrane perturbation protocol to efficiently screen receptor candidates involved in hetero-multivalent protein binding.

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Section: Discussionmentioning

confidence: 99%

Hetero-multivalent binding of cholera toxin subunit B with glycolipid mixtures

Krishnan

Singla

Lee

et al. 2017

Colloids and Surfaces B: Biointerfaces

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“…For example, interactions between fucosylated glycans (such as blood group antigens) and bacterial cholesterol-dependent pore forming toxins (e.g, pneumolysin and streptolysin O) promote insertion of these toxins within the host cell membrane, and binding to fucosylated glycans appears to promote host cell uptake of cholera toxin (Shewell et al, 2014; Wands et al, 2015). Thus, fucosylated surface molecules may serve as receptors for T3SS2 translocon components, whose binding may be an obligatory step in translocon insertion.…”

Section: Discussionmentioning

confidence: 99%

CRISPR/Cas9 Screens Reveal Requirements for Host Cell Sulfation and Fucosylation in Bacterial Type III Secretion System-Mediated Cytotoxicity

Blondel

Park

Hubbard

et al. 2016

Cell Host & Microbe

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SUMMARY Type III secretion systems (T3SSs) inject bacterial effector proteins into host cells and underlie the virulence of many Gram-negative pathogens. Studies have illuminated bacterial factors required for T3SS function, but the required host processes remain largely undefined. We coupled CRISPR/Cas9 genome editing technology with the cytotoxicity of two Vibrio parahaemolyticus T3SSs (T3SS1 and T3SS2) to identify human genome disruptions conferring resistance to T3SS-dependent cytotoxicity. We identity non-overlapping genes required for T3SS1- and T3SS2-mediated cytotoxicity. Genetic ablation of cell surface sulfation reduces bacterial adhesion and thereby alters the kinetics of T3SS1-mediated cytotoxicity. Cell surface fucosylation is required for T3SS2-dependent killing, and pharmacological or genetic inhibition of fucosylation prevents membrane insertion of the T3SS2 translocon complex. These findings reveal the importance of ubiquitous surface modifications for T3SS function, potentially explaining the broad tropism of V. parahaemolyticus, as well as highlight the utility of genome-wide CRISPR/Cas9 screens to discover processes underlying host-pathogen interactions.

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“…First, affinity labels often cannot access the entire cell surface due to their relatively large size; second, lipids that are already bound to endogenous proteins cannot be detected; third, affinity label binding often depends on the specific orientation and/or clustering of the target lipid (Mahfoud et al, 2010; Mizuno et al, 2011; Kishimoto et al, 2016). Another disadvantage is that some anti-glycosphingolipid antibodies and the popular affinity label for GM1, cholera toxin subunit B, may also bind to glycoproteins, which compromises their ability to report the distribution of the target glycosphingolipid (Tonegawa and Hakomori, 1977; Blank et al, 2007; Day and Kenworthy, 2012; Wands et al, 2015). …”

Section: Methods To Image Sphingolipid Distribution In the Plasma Memmentioning

confidence: 99%

Sphingolipid Organization in the Plasma Membrane and the Mechanisms That Influence It

Kraft

2017

Front. Cell Dev. Biol.

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Sphingolipids are structural components in the plasma membranes of eukaryotic cells. Their metabolism produces bioactive signaling molecules that modulate fundamental cellular processes. The segregation of sphingolipids into distinct membrane domains is likely essential for cellular function. This review presents the early studies of sphingolipid distribution in the plasma membranes of mammalian cells that shaped the most popular current model of plasma membrane organization. The results of traditional imaging studies of sphingolipid distribution in stimulated and resting cells are described. These data are compared with recent results obtained with advanced imaging techniques, including super-resolution fluorescence detection and high-resolution secondary ion mass spectrometry (SIMS). Emphasis is placed on the new insight into the sphingolipid organization within the plasma membrane that has resulted from the direct imaging of stable isotope-labeled lipids in actual cell membranes with high-resolution SIMS. Super-resolution fluorescence techniques have recently revealed the biophysical behaviors of sphingolipids and the unhindered diffusion of cholesterol analogs in the membranes of living cells are ultimately in contrast to the prevailing hypothetical model of plasma membrane organization. High-resolution SIMS studies also conflicted with the prevailing hypothesis, showing sphingolipids are concentrated in micrometer-scale membrane domains, but cholesterol is evenly distributed within the plasma membrane. Reductions in cellular cholesterol decreased the number of sphingolipid domains in the plasma membrane, whereas disruption of the cytoskeleton eliminated them. In addition, hemagglutinin, a transmembrane protein that is thought to be a putative raft marker, did not cluster within sphingolipid-enriched regions in the plasma membrane. Thus, sphingolipid distribution in the plasma membrane is dependent on the cytoskeleton, but not on favorable interactions with cholesterol or hemagglutinin. The alternate views of plasma membrane organization suggested by these findings are discussed.

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Fucosylation and protein glycosylation create functional receptors for cholera toxin

Cited by 88 publications

References 72 publications

Hetero-multivalent binding of cholera toxin subunit B with glycolipid mixtures

Hetero-multivalent binding of cholera toxin subunit B with glycolipid mixtures

CRISPR/Cas9 Screens Reveal Requirements for Host Cell Sulfation and Fucosylation in Bacterial Type III Secretion System-Mediated Cytotoxicity

Sphingolipid Organization in the Plasma Membrane and the Mechanisms That Influence It

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