The enzyme subunit SubA of Shiga toxin-producing E. coli strains demonstrates comparable intracellular transport and cytotoxic activity as the holotoxin SubAB in HeLa and HCT116 cells in vitro
Copyright policy )The enzyme subunit SubA of Shiga toxin-producing E. coli strains demonstrates comparable intracellular transport and cytotoxic activity as the holotoxin SubAB in HeLa and HCT116 cells in vitro
AbstractThe subtilase cytotoxin (SubAB) is secreted by certain Shiga toxin-producing Escherichia coli (STEC) strains and is composed of the enzymatically active subunit SubA and the pentameric binding/transport subunit SubB. We previously demonstrated that SubA (10 µg/ml), in the absence of SubB, binds and intoxicates the human cervix cancer-derived epithelial cell line HeLa. However, the cellular and molecular mechanisms underlying the cytotoxic activity of SubA in the absence of SubB remained unclear. In the present study, the cytotoxic effects mediated by SubA alone were investigated in more detail in HeLa cells and the human colon cancer cell line HCT116. We found that in the absence of SubB, SubA (10 µg/ml) is internalized into the endoplasmic reticulum (ER), where it cleaves the chaperone GRP78, an already known substrate for SubA after its canonical uptake into cells via SubB. The autonomous cellular uptake of SubA and subsequent cleavage of GRP78 in cells is prevented by treatment of cells with 10 µM brefeldin A, which inhibits the transport of protein toxins into the ER. In addition, by analyzing the SubA mutant SubAΔC344, we identified the C-terminal SEEL motif as an ER-targeting signal. Conclusively, our results strongly suggest that SubA alone shares the same intracellular transport route and cytotoxic activity as the SubAB holotoxin.
- University of Ulm Germany
- University of Hohenheim Germany
- Institut für Pharmakologie und Toxikologie der Bundeswehr Germany
GRP78, Molecular chaperones, Uterine Cervical Neoplasms, Subtilase cytotoxin, Intrazellulärer Transport, Molecular Toxicology, Cellular uptake, Escherichia coli, Humans, Glycosides, Subtilisins, Endoplasmic Reticulum Chaperone BiP, Heat-Shock Proteins, info:eu-repo/classification/ddc/610, Shiga-Toxigenic Escherichia coli, Shiga toxin-producing Escherichia coli (STEC), Hitzeschock-Proteine, Escherichia coli Proteins, Biological Transport, HCT116 Cells, Triterpenes, STEC, Shiga-toxigenic Escherichia coli, Colonic Neoplasms, Shiga toxins, Heat-shock proteins, Female, Escherichia coli Proteins/metabolism [MeSH] ; Glycosides/pharmacology [MeSH] ; Subtilase cytotoxin ; Uterine Cervical Neoplasms/pathology [MeSH] ; Triterpenes/pharmacology [MeSH] ; Colonic Neoplasms/pathology [MeSH] ; HCT116 Cells [MeSH] ; Subtilisins/metabolism [MeSH] ; Biological Transport [MeSH] ; Subtilisins/pharmacology [MeSH] ; Female [MeSH] ; Escherichia coli Proteins/pharmacology [MeSH] ; Heat-Shock Proteins/metabolism [MeSH] ; Molecular Toxicology ; Humans [MeSH] ; GRP78 ; Uterine Cervical Neoplasms/drug therapy [MeSH] ; Shiga toxin-producing ; Intracellular transport ; Cellular uptake ; Colonic Neoplasms/drug therapy [MeSH] ; Shiga-Toxigenic Escherichia coli/metabolism [MeSH] ; Triterpenes/metabolism [MeSH] ; HeLa Cells [MeSH] ; Glycosides/metabolism [MeSH], Intracellular transport, HeLa Cells
GRP78, Molecular chaperones, Uterine Cervical Neoplasms, Subtilase cytotoxin, Intrazellulärer Transport, Molecular Toxicology, Cellular uptake, Escherichia coli, Humans, Glycosides, Subtilisins, Endoplasmic Reticulum Chaperone BiP, Heat-Shock Proteins, info:eu-repo/classification/ddc/610, Shiga-Toxigenic Escherichia coli, Shiga toxin-producing Escherichia coli (STEC), Hitzeschock-Proteine, Escherichia coli Proteins, Biological Transport, HCT116 Cells, Triterpenes, STEC, Shiga-toxigenic Escherichia coli, Colonic Neoplasms, Shiga toxins, Heat-shock proteins, Female, Escherichia coli Proteins/metabolism [MeSH] ; Glycosides/pharmacology [MeSH] ; Subtilase cytotoxin ; Uterine Cervical Neoplasms/pathology [MeSH] ; Triterpenes/pharmacology [MeSH] ; Colonic Neoplasms/pathology [MeSH] ; HCT116 Cells [MeSH] ; Subtilisins/metabolism [MeSH] ; Biological Transport [MeSH] ; Subtilisins/pharmacology [MeSH] ; Female [MeSH] ; Escherichia coli Proteins/pharmacology [MeSH] ; Heat-Shock Proteins/metabolism [MeSH] ; Molecular Toxicology ; Humans [MeSH] ; GRP78 ; Uterine Cervical Neoplasms/drug therapy [MeSH] ; Shiga toxin-producing ; Intracellular transport ; Cellular uptake ; Colonic Neoplasms/drug therapy [MeSH] ; Shiga-Toxigenic Escherichia coli/metabolism [MeSH] ; Triterpenes/metabolism [MeSH] ; HeLa Cells [MeSH] ; Glycosides/metabolism [MeSH], Intracellular transport, HeLa Cells
selected citations These citations are derived from selected sources.This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).5 popularity This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.Top 10% influence This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).Average impulse This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.Average
Citations provided by BIP!Popularity provided by BIP!