Dual infection system identifies a crucial role for PKA-mediated serine phosphorylation of the EPEC-Tir-injected effector protein in regulating Rac1 function.
Many Gram-negative pathogenic bacteria possess type-III or type-IV secretion systems to inject 'effector' proteins directly into host cells to modulate cellular processes in their favour. A common target is the actin-cytoskeleton linked to the delivery of a single (CagA) effector by Helicobacter pylori and multiple effectors by enteropathogenic Escherichia coli (EPEC) respectively. Here we report co-infection as a powerful strategy for defining effector protein function and mechanisms by which they modulate cellular responses. This is exemplified by our finding that EPEC inhibits H. pylori-induced AGS cell elongation, a disease-related event linked to Rac1 activation. While this inhibitory process is dependent on the translocated Intimin receptor, Tir, and the outer-membrane protein, Intimin, it unexpectedly revealed evidence for Tir signalling independent of Intimin interaction and tyrosine phosphorylation of Tir. Furthermore, the work defined a long awaited role for protein kinase A (PKA)-mediated phosphorylation of Tir at serine-434 and serine-463. Our data are consistent with a model whereby EPEC activates PKA for Tir phosphorylation. Activated PKA then phosphorylates Rac1 at serine-71 associated with reduced GTP-load and inhibited cell elongation. Thus, the co-infection approach is a powerful strategy for defining novel effector function with important implications for characterizing mechanisms and regulatory signalling pathways in bacterial pathogenesis.
Brandt
Sabine
S
Department of Medical Microbiology, Otto von Guericke University, Leipziger Str. 44, D-39120 Magdeburg, Germany.
Kenny
Brendan
B
Rohde
Manfred
M
Martinez-Quiles
Narcisa
N
Backert
Steffen
S
D018829:Adhesins, Bacterial / Q000378:metabolism; D000942:Antigens, Bacterial / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D001427:Bacterial Toxins / Q000378:metabolism; D045744:Cell Line, Tumor; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D054308:Enteropathogenic Escherichia coli / Q000378:metabolism* / Q000472:pathogenicity; D004927:Escherichia coli Infections / Q000150:complications*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D016481:Helicobacter Infections / Q000150:complications*; D016480:Helicobacter pylori / Q000378:metabolism / Q000472:pathogenicity; D054884:Host-Pathogen Interactions; D006801:Humans; D010766:Phosphorylation; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism*; D012694:Serine / Q000737:chemistry / Q000378:metabolism*; D014774:Virulence; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
D018829:Adhesins, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D029968:Escherichia coli Proteins; C107953:RAC1 protein, human; D011956:Receptors, Cell Surface; C109287:Tir protein, E coli; C080866:cagA protein, Helicobacter pylori; C060296:cytotoxic necrotizing factor type 1; C073819:eaeA protein, E coli; D012694:Serine; D017868:Cyclic AMP-Dependent Protein Kinases; D020830:rac1 GTP-Binding Protein