A low-Ca2+ response operon encodes the V antigen of Yersinia pestis

Abstract

Yersinia pestis has a virulence regulon called the low-Ca2+ response that is mediated by the plasmid pCD and manifested as regulation of growth and of expression of several virulence-associated properties by Ca2+ and temperature. We used Mu dI(Ap lac) to obtain a mutation in pCD1 of Y. pestis KIM that rendered the bacteria unable to express one of these properties, the V antigen. This mutant also had lost the Ca2+ requirement for growth at 37 degrees C and was avirulent in mice. Two-dimensional protein gel electrophoresis showed that the Mu dI(Ap lac) insertion had eliminated 13,000- and 18,000-molecular-weight proteins in addition to the V antigen. We mapped the Mu dI(Ap lac) insertion within pCD1, cloned the HindIII fragment spanning the insertion location, prepared two subclones of this fragment, and identified the proteins these clones expressed in Escherichia coli minicells. The data indicated that the V gene lies within an operon containing three genes; lcrG (encoding the 13,000-molecular-weight protein), lcrV (encoding the 38,000-molecular-weight V antigen), and lcrH (encoding the 18,000-molecular-weight protein). Therefore, the V operon contains the structural gene for V antigen, at least one virulence gene, and at least one Ca2+-dependence gene.