FIG. 1.

lcrG mutant yersiniae secrete Yops in the presence of calcium. Y. enterocolitica W22703 (wild type) and the isogenic lcrG mutant, strain MC2, were transformed with pDA37 (vector control) or pCT60 (wild-type lcrG) and grown in the presence of 5 mM calcium or 5 mM EGTA for 2 h at 37°C. Cultures were centrifuged, and the supernatant (S) was separated from the cell pellet (P). Proteins in each sample were precipitated with TCA, solubilized in sample buffer, and analyzed by SDS-PAGE, followed by immunoblotting with antisera raised against LcrG, LcrV, LcrH, YopB, YopD, YopE, and CAT. Y. enterocolitica W22703 secretes more YopBDE in the absence than in the presence of calcium. In contrast, the lcrG mutant strain displays a calcium-blind phenotype and secretes YopBDE in the presence or absence of calcium. Further, the lcrG mutant failed to secrete LcrV. Transformation of lcrG mutant cells with pCT60 restored the wild-type phenotype.