Uptake of Shiga-toxigenic Escherichia coliâ
SubAB by HeLa cells requires an actin- and lipid raft-dependent pathway.
Cell Microbiol
; 16(10): 1582-601, 2014 Oct.
Article
em En
| MEDLINE
| ID: mdl-24844382
ABSTRACT
The novel cytotoxic factor subtilase cytotoxin (SubAB) is produced mainly by non-O157 Shiga-toxigenic Escherichia coli (STEC). SubAB cleaves the molecular chaperone BiP/GRP78 in the endoplasmic reticulum (ER), leading to activation of RNA-dependent protein kinase (PKR)-like ER kinase (PERK), followed by caspase-dependent cell death. However, the SubAB uptake mechanism in HeLa cells is unknown. In this study, a variety of inhibitors and siRNAs were employed to characterize the SubAB uptake process. SubAB-induced BiP cleavage was inhibited by high concentrations of Dynasore, and methyl-ß-cyclodextrin (mßCD) and Filipin III, but not suppressed in clathrin-, dynamin I/II-, caveolin1- and caveolin2-knockdown cells. We observed that SubAB treatment led to dramatic actin rearrangements, e.g. formation of plasma membrane blebs, with a significant increase in fluid uptake. Confocal microscopy analysis showed that SubAB uptake required actin cytoskeleton remodelling and lipid raft cholesterol. Furthermore, internalized SubAB in cells was found in the detergent-resistant domain (DRM) structure. Interestingly, IPA-3, an inhibitor of serine/threonine kinase p21-activated kinase (PAK1), an important protein of macropinocytosis, directly inhibited SubAB-mediated BiP cleavage and SubAB internalization. Thus, our findings suggest that SubAB uses lipid raft- and actin-dependent, but not clathrin-, caveolin- and dynamin-dependent pathways as its major endocytic translocation route.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Citoesqueleto de Actina
/
Subtilisinas
/
Microdomínios da Membrana
/
Proteínas de Escherichia coli
/
Escherichia coli Shiga Toxigênica
/
Proteínas de Choque Térmico
Limite:
Humans
Idioma:
En
Revista:
Cell Microbiol
Assunto da revista:
MICROBIOLOGIA
Ano de publicação:
2014
Tipo de documento:
Article
País de afiliação:
Japão