Yersinia enterocolitica TyeA, an intracellular regulator of the type II machinery, is required for specific targeting of YopE, YopH, YopM, and YopN into the cytosol of eukaryotic cells

Pathogenic Yersinia species employ type III machines to target effector Yop s into the cytosol of eukaryotic cells. Yersinia tyeA mutants are thought t o be defective in the targeting of YopE and YopH without affecting the inje ction of YopM, YopN, YopO, YopP, and YopT into the cytosol of eukaryotic ce lls. One model suggests that TyeA may form a tether between YopN (LcrE) and YopD on the bacterial surface, a structure that may translocate YopE and Y opH across the plasma membrane of eukaryotic cells (NI. Iriarte, M. P. Sory , A. Poland, A. P. Boyd, S. D. Mills, I. Lambermont, and G. R. Cornelis, EM PO J. 17:1907-1918, 1998). We have examined the injection of Yop proteins b y tyeA mutant yersiniae with the digitonin fractionation technique. We find that tyeA mutant yersiniae not only secreted YopE, YopH, YopM, and YopN in to the extracellular medium but also targeted these polypeptides into the c ytosol of HeLa cells. Protease protection, cell fractionation, and affinity purification experiments suggest that TyeA is located intracellularly and binds to YopN or YopD. We propose a model whereby TyeA functions as a negat ive regulator of the type III targeting pathway in the cytoplasm of yersini ae, presumably by preventing the export of YopN.