Enteropathogenic E. coli Tir binds Nck to initiate actin pedestal formation in host cells.
Enteropathogenic Escherichia coli (EPEC) is a bacterial pathogen that causes infantile diarrhea worldwide. EPEC injects a bacterial protein, translocated intimin receptor (Tir), into the host-cell plasma membrane where it acts as a receptor for the bacterial outer membrane protein, intimin. The interaction of Tir and intimin triggers a marked rearrangement of the host actin cytoskeleton into pedestals beneath adherent bacteria. On delivery into host cells, EPEC Tir is phosphorylated on tyrosine 474 of the intracellular carboxy-terminal domain, an event that is required for pedestal formation. Despite its essential role, the function of Tir tyrosine phosphorylation has not yet been elucidated. Here we show that tyrosine 474 of Tir directly binds the host-cell adaptor protein Nck, and that Nck is required for the recruitment of both neural Wiskott-Aldrich-syndrome protein (N-WASP) and the actin-related protein (Arp)2/3 complex to the EPEC pedestal, directly linking Tir to the cytoskeleton. Cells with null alleles of both mammalian Nck genes are resistant to the effects of EPEC on the actin cytoskeleton. These results implicate Nck adaptors as host-cell determinants of EPEC virulence.
Gruenheid
S
S
Biotechnology Laboratory, University of British Columbia, 6174 University Boulevard, Vancouver V6T 1G3, Canada.
DeVinney
R
R
Bladt
F
F
Goosney
D
D
Gelkop
S
S
Gish
G D
GD
Pawson
T
T
Finlay
B B
BB
D000199:Actins / Q000378:metabolism*; D018829:Adhesins, Bacterial; D000595:Amino Acid Sequence; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000502:physiology; D001426:Bacterial Proteins / Q000378:metabolism; D001665:Binding Sites; D002352:Carrier Proteins; D002460:Cell Line; D002462:Cell Membrane / Q000382:microbiology* / Q000502:physiology / Q000648:ultrastructure; D003599:Cytoskeleton / Q000502:physiology / Q000648:ultrastructure; D019453:Escherichia coli O157 / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*; D029968:Escherichia coli Proteins; D005347:Fibroblasts / Q000166:cytology / Q000382:microbiology / Q000502:physiology / Q000648:ultrastructure; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D006801:Humans; D007700:Kinetics; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics; D008322:Mammals; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000032:analysis; D014162:Transfection; D018909:src Homology Domains
D000199:Actins; D018829:Adhesins, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D008164:Luminescent Proteins; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; C109287:Tir protein, E coli; C073819:eaeA protein, E coli; D049452:Green Fluorescent Proteins; D019000:Phosphotyrosine