Modulation of the low-calcium response in Yersinia pestis via plasmid-plasmid interaction

Virulent cells of Yersinia pestis, Yersinia pseudotuberculosis, and Yersinia enterocolitica are known to exhibit a low-calcium response in vitro characterized by restriction of growth and induction of V antigen at 37 degrees C in Ca2+-deficient media (Lcr+). A shared Lcr plasmid mediates these properties and encodes yersiniae outer membrane peptides (Yops) that are expressed in Lcr+ Y. pseudotuberculosis and Y. enterocolitica but not Y. pestis. We present direct evidence here verifying that synthesis of major Yops in the former two species is, like V, repressed by Ca2+ and that these structures are located primarily in the outer membrane. We also verified that rabbits infected with live Lcr+ Y. pestis can raise antibodies against V and Yops. Similar antisera, however, were recovered after immunization with sterile extracts of Ca2+-starved Lcr+ cells of Y. pestis. Results of immunoblots obtained with these antisera showed that restricted Y. pestis produced Yops of about 46 kDa (YopB) and 44 kDa (YopC) after shiftup by addition of Ca2+ at 37 degrees C or reduction of temperature to 26 degrees C. It is established that virulent cells of Y. pestis also possess a unique plasmid known to mediate pesticinogeny (Pst+). Restricted Lcr+, Pst- Y. pestis expressed YopB and YopC plus additional 76 kDa (YopF), 48 kDa (YopH), 36 kDa (YopD), 32.5 kDa (YopJ), and 27 kDa (YopE) outer membrane structures at concentrations comparable to those in Ca2+-starved Y. pseudotuberculosis and Y. enterocolitica. These findings indicate that carriage of the Pst plasmid prevents expression of the Lcr plasmid-mediated Yops in wild type Y. pestis.