RfaL Is Required for Yersinia pestis Type III Secretion and Virulence

Abstract

ABSTRACTYersinia pestis, the causative agent of plague, uses a type III secretion system (T3SS) to inject cytotoxic Yop proteins directly into the cytosol of mammalian host cells. The T3SS can also be activatedin vitroat 37°C in the absence of calcium. The chromosomal generfaL(waaL) was recently identified as a virulence factor required for proper function of the T3SS. RfaL functions as a ligase that adds the terminalN-acetylglucosamine to the lipooligosaccharide core ofY. pestis. We previously showed that deletion ofrfaLprevents secretion of Yopsin vitro. Here we show that the divalent cations calcium, strontium, and magnesium can partially or fully rescue Yop secretionin vitro, indicating that the secretion phenotype of therfaLmutant may be due to structural changes in the outer membrane and the corresponding feedback inhibition on the T3SS. In support of this, we found that the defect can be overcome by deleting the regulatory genelcrQ. Consistent with a defective T3SS, therfaLmutant is less virulent than the wild type. We show here that the virulence defect of the mutant correlates with a decrease in both T3SS gene expression and ability to inject innate immune cells, combined with an increased sensitivity to cationic antimicrobial peptides.