THE YERSINIA YPKA SER THR KINASE IS TRANSLOCATED AND SUBSEQUENTLY TARGETED TO THE INNER SURFACE OF THE HELA-CELL PLASMA-MEMBRANE/

Multiple yop mutant strains of Yersinia pseudotuberculosis not express ing several virulence effector Yop proteins (YopH, M, E, K and YpkA) w ere engineered. When high-copy-number plasmids carrying the ypkA or th e yopE gene with their endogenous promoters were introduced into the e ngineered strains, the corresponding Yop protein was secreted at high levels in vitro. These multiple yop mutant strains, when harbouring th e yopE gene in trans, behaved as the wild-type strain with respect to YopB-dependent translocation of YopE through the HeLa cell plasma memb rane. Using these multiple yop mutant strains, it was demonstrated tha t the YpkA Ser/Thr protein kinase mediates morphological alterations o f infected cultured HeLa cells different from those mediated by YopE a nti YopH. Furthermore, YpkA is shown to be translocated by a YopB-depe ndent translocation mechanism from surface-located bacteria and subseq uently targeted to the inner surface of the target-cell plasma membran e. The pattern of YpkA localization after infection suggests that this Yop effector is involved In interference with signal transduction.