A hallmark of Yersinia type III machines is the presence of needles extending from the bacterial surface. Needles perform two functions, serving as the conduits for the transport of effectors into immune cells but also acting as a sensor. The polymerized needle protein, YscF, is thought to perceive threshold levels of environmental calcium ions to trigger secretion. yopR (yscH) is a gene downstream of yscEFG, encoding the chaperones and principal building blocks of the needle. Here we investigated the contribution of YopR towards type III secretion and pathogenesis. Yersinia pestis KIM D27 mutants lacking yopR were defective for virulence in a mouse model of septicemic plague. yopR variants of Yersinia enterocolitica W22703 displayed a reduced ability to inject effectors into macrophages and required lower calcium concentrations to activate type III secretion than wild-type yersiniae. Furthermore, yopR mutants failed to assemble YscF into needle complexes and instead secreted YscF into the medium. These results imply that YopR may be involved in controlling the secretion of YscF, thereby impacting the assembly of type III machines. An alternative possibility, which YopR participates directly in the polymerization of YscF, seems less likely as YopR is not associated with purified needles.