Enteropathogenic Escherichia coli Tir translocation and pedestal formation requires membrane cholesterol in the absence of bundle-forming pili.
Enteropathogenic Escherichia coli (EPEC) is a significant cause of paediatric diarrhoea worldwide. Virulence requires adherence to intestinal epithelial cells, mediated in part through type IV bundle-forming pili (BFP), and the EPEC protein Tir. Tir is inserted into the enterocyte plasma membrane (PM), resulting in the formation of actin-rich pedestals. Tir is translocated by the type III secretion system (TTSS), through a pore comprised of EPEC proteins inserted into the PM. Here, we demonstrate that in the absence of BFP, EPEC adherence, effector translocation and pedestal formation are dependent on lipid rafts. Lipid raft disruption using methyl-beta-cyclodextrin (MbetaCD) decreased adherence by an EPEC BFP-deficient strain from 85% to 1%. Translocation of the effectors Tir and EspF was blocked by MbetaCD treatment, although the TTSS pore still formed. MbetaCD treatment after Tir delivery decreased pedestal formation by EPEC from 40% to 5%, but not by the related pathogen E. coli O157:H7 which uses a different Tir-based mechanism. In contrast, EPEC expressing the BFP can circumvent the requirement for membrane cholesterol. This suggests that lipid rafts play a role in virulence of this medically important pathogen.
University of Calgary Health Sciences Centre, Department of Microbiology and Infectious Diseases, 3330 Hospital Drive NW, Calgary, Alberta T2N 4N1, Canada.
Emma
E
Allen-Vercoe
Barbara
B
Waddell
Scott
S
Livingstone
Julie
J
Deans
Rebekah
R
DeVinney
D001422:Bacterial Adhesion; D002352:Carrier Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000502:physiology*; D002784:Cholesterol / Q000502:physiology*; D003967:Diarrhea / Q000382:microbiology; D020895:Enterocytes / Q000382:microbiology / Q000502:physiology; D004926:Escherichia coli / Q000502:physiology*; D004927:Escherichia coli Infections / Q000382:microbiology; D029968:Escherichia coli Proteins / Q000378:metabolism / Q000502:physiology*; D010861:Fimbriae, Bacterial / Q000502:physiology*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D021962:Membrane Microdomains / Q000502:physiology; D021381:Protein Transport; D011956:Receptors, Cell Surface / Q000502:physiology*; D047392:beta-Cyclodextrins / Q000494:pharmacology
D002352:Carrier Proteins; D029968:Escherichia coli Proteins; C489109:EspFU protein, E coli; D047908:Intracellular Signaling Peptides and Proteins; D011956:Receptors, Cell Surface; C109287:Tir protein, E coli; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D002784:Cholesterol