Enteropathogenic Escherichia coli (EPEC) Tir receptor molecule does not undergo full modification when introduced into host cells by EPEC-independentmechanisms

Enteropathogenic Escherichia coli (EPEC), like many other gram-negative pat hogens, encodes a type III secretion apparatus dedicated to the release of virulence-associated proteins. One such protein, Tir, is translocated into host cells, where it is modified by the addition of phosphate groups, resul ting in a number of species with distinct molecular mass. One phosphorylati on event, on tyrosine residue 474 of Tir, does not contribute to shifts in molecular mass but is essential for its actin-nucleating function. The role of the nonphosphotyrosine related modifications is unknown. In this paper, we demonstrate, using three different approaches, that Tir does not encode sufficient information to facilitate its complete modification when introd uced into host cells in EPEC-independent mechanisms. Each system revealed t hat Tir is a substrate for a host kinase whose action results in its partia l modification to a form similar to one evident in EPEC-infected host cells . Further Tir modification could not be induced by infecting cells with EPE C, suggesting that Tir must be coexpressed with other EPEC factors to enabl e its full modification within host cells. One approach used Yersinia spp. to deliver Tir into host cells, and this system revealed that Tir secretion and translocation can occur in the absence of the Tir chaperone molecule, CesT (formerly known as OrfU). CesT was found to be an efficiency factor wh ich was not required, unlike in EPEC, for Tir stability, indicating that it may function to guide Tir to the translocation apparatus or maintain it in a secretion-competent form.