Proteoglycan-Dependent Endo-Lysosomal Fusion Affects Intracellular Survival of Salmonella Typhimurium in Epithelial Cells

Abstract

Proteoglycans (PGs) are glycoconjugates which are predominately expressed on cell surfaces and consist of glycosaminoglycans (GAGs) linked to a core protein. An initial step of GAGs assembly is governed by the β-D-xylosyltransferase enzymes encoded in mammals by the XylT1/XylT2 genes. PGs are essential for the interaction of a cell with other cells as well as with the extracellular matrix. A number of studies highlighted a role of PGs in bacterial adhesion, invasion, and immune response. In this work, we investigated a role of PGs in Salmonella enterica serovar Typhimurium (S. Typhimurium) infection of epithelial cells. Gentamicin protection and chloroquine resistance assays were applied to assess invasion and replication of S. Typhimurium in wild-type and xylosyltransferase-deficient (XylT2) Chinese hamster ovary (CHO) cells lacking PGs. We found that S. Typhimurium adheres to and invades CHO WT and CHO XylT2 cells at comparable levels. However, 24 h after infection, proteoglycandeficient CHO XylT2 cells are significantly less colonized by S. Typhimurium compared to CHO WT cells. This proteoglycan-dependent phenotype could be rescued by addition of PGs to the cell culture medium, as well as by complementation of the XylT2 gene. Chloroquine resistance assay and immunostaining revealed that in the absence of PGs, significantly less bacteria are associated with Salmonella-containing vacuoles (SCVs) due to a redistribution of endocytosed gentamicin. Inhibition of endolysosomal fusion by a specific inhibitor of phosphatidylinositol phosphate kinase PIKfyve significantly increased S. Typhimurium burden in CHO XylT2 cells demonstrating an important role of PGs for PIKfyve dependent vesicle fusion which is modulated by Salmonella to establish infection. Overall, our results demonstrate that PGs influence survival of intracellular Salmonella in epithelial cells via modulation of PIKfyve-dependent endo-lysosomal fusion.

Limosilactobacillus reuteri SLZX19-12 Protects the Colon from Infection by Enhancing Stability of the Gut Microbiota and Barrier Integrity and Reducing Inflammation

“…Surprisingly, the expression of the barrier proteins occludin, claudin-1, claudin-2, claudin-7, and EpCAM was not significantly influenced by S. Typhimurium SL1344, as was reported by Sun et al ( 40 ). According to previous research, Salmonella usually breaks through intestinal epithelial cells to the lamina propria of the mucosa via transcytosis of M cells, epithelial endocytosis, and phagocyte uptake ( 41 – 43 ). In the colon, virulence proteins from Salmonella interact with ordinary intestinal epithelial cells and rearrange the cytoskeleton ( 44 ).…”

Bright individuals: Applications of fluorescent protein‐based reporter systems in single‐cell cellular microbiology

“…As reporters in the bacterial cytosol, FPs expressed by DFR are less likely to be affected by pH changes due to the pH homeostasis in the bacterial cytosol. However, extreme external conditions such as presence in phagolysosome of host cells can result in acid shock and override pH homeostasis, as determined by DFR with an acid shock‐induced promoter (Galeev et al., 2020). Although drop of cytosolic pH will be transient due to compensation by acid shock response, pH‐mediated effects on FP signal intensities should be considered in interpretation of results.…”

Strategies and Future Opportunities for the Prevention, Diagnosis, and Management of Cow Milk Allergy

“…A component of balancing the response occurs in the 3rd trimester through allergen aggregated with IgG transported across the placenta which down-regulates sensitization through inhibitory IgG receptors. This being the case, high maternal intake of milk would increase IgG antibodies complexed with milk and when transported across the placenta could reduce the risk of CMA ( 23 ). Balancing the Th-1 and Th-2 responses after delivery is affected by a rapidly diversifying ecological balance of organisms (microbiome) on the infant’s skin, in the respiratory and gastrointestinal tracts.…”

“…In animal models, it has been shown that antigen-immunoglobulin (IgG) immune complexes from sensitized dams were transferred to the newborn through the neonatal Fc receptor resulting in the induction of antigen-specific FoxP3(+) CD25(+) regulatory T cells ( 43 ). The transfer of these immune complexes could induce tolerance and support the development of primary prevention of CMA ( 23 , 44 ).…”