Abstract
Shiga toxin (Stx) is an enterotoxin produced by Shigella dysenteriae serotype 1 and enterohemorrhagic Escherichia coli, which binds specifically to globotriaosylceramide, Gb3, on the cell surface and causes cell death. We previously demonstrated that Stx induced apoptosis in human renal tubular cell line ACHN cells (Taguchi, T., Uchida, H., Kiyokawa, N., Mori, T., Sato, N., Horie, H., Takeda, T and Fujimoto, J. (1998) Kidney Int. 53, 1681-1688). To study the early signal transduction after Stx addition, Gb3-enriched microdomains were prepared from ACHN cells by sucrose density gradient centrifugation of Triton X-100 lysate as buoyant, detergent-insoluble microdomains (DIM). Gb3 was only recovered in DIM and was associated with Src family kinase Yes. Phosphorylation of tyrosine residues of proteins in the DIM fraction increased by 10 min and returned to the resting level by 30 min after the addition of Stx. Since the kinase activity of Yes changed with the same kinetics, Yes was thought to be responsible for the hyperphosphorylation observed in DIM proteins. Unexpectedly, however, all of the Yes kinase activity was obtained in the high density, detergent-soluble fraction. Yes was assumed to be activated and show increased Triton X-100 solubility in the early phase of retrograde endocytosis of Stx-Gb3 complex. Since Yes activation by the Stx addition was suppressed by filipin pretreatment, Gb3-enriched microdomains containing cholesterol were deeply involved in Stx signal transduction.
Highlights
Cell death is widely known to take place through two distinctive processes, necrosis or apoptosis
We previously demonstrated that Shiga toxin (Stx) induced apoptosis in human renal tubular cell line ACHN cells (Taguchi, T., Uchida, H., Kiyokawa, N., Mori, T., Sato, N., Horie, H., Takeda, T and Fujimoto, J. (1998) Kidney Int. 53, 1681–1688)
In contradiction to Williams’s report [4], a number of recent studies have clearly demonstrated that Stx induces apoptosis in several different cell types, including Burkitt’s lymphoma cells [5], Vero cells [6], human renal tubular derived ACHN cells [1], and normal human renal tubular epithelial cells [7, 8]
Summary
Shiga toxin; HUS, hemolytic uremic syndrome; GSL, glycosphingolipid; mAb, monoclonal antibody; DIM, detergent-insoluble microdomain(s); GPI, glycosylphosphatidylienterohemorrhagic Escherichia coli is one of the major cause of hemolytic uremic syndrome (HUS). The B subunit has no inhibitory effect on protein synthesis, a series of studies indicates that it alone participate to transduce cell signaling and can induce apoptosis in some instances such as Burkitt’s lymphoma [5, 9]. These reports encourage us to determine how and where Gb3 delivers the signal. Some gangliosides in DIM were reported to be associated with transducer molecules that are activated by stimulation of ganglioside [11, 12] By analogy to this model, the Stx receptor Gb3 is expected to reside in DIM and to be involved in signal transduction. The activation-related topology of Yes and the recruitment of Yes after Stx addition are discussed
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