TLR2-Mediated Signaling Requirements for Francisella tularensis LVS Infection of Murine Macrophages. [Infect Immun] Journal article

Francisella tularensis (Ft), an aerobic, non-spore forming, Gram-negative coccobaccillus, is the causative agent of tularemia. We reported previously that Ft Live Vaccine Strain (LVS) elicited strong, dose-dependent NF-kappaB reporter activity in TLR2-expressing HEK293T cells and pro-inflammatory gene expression in primary murine macrophages. Herein, we report that Ft LVS-induced murine macrophage pro-inflammatory cytokine gene and protein expression is overwhelmingly TLR2-dependent as evidenced by the abrogated responses of TLR2(-/-) macrophages. Ft LVS infection also increased expression of TLR2 both in vitro, in mouse macrophages, and in vivo, in livers from Ft LVS-infected mice. Colocalization of intracellular Ft LVS, TLR2, and MyD88 was visualized by confocal microscopy. Signaling was abrogated if the Ft LVS were heat- or formalin-killed or treated with chloramphenicol, indicating that the TLR2 agonist activity is dependent on new bacterial protein synthesis. Ft LVS replicates in macrophages; however, bacterial replication was not required for TLR2 signaling because LVSDeltaguaA, a Ft LVS guanine auxotroph that fails to replicate in the absence of exogenous guanine, activated NF-kappaB in TLR2-transfected HEK293T cells and induced cytokine expression in wild-type macrophages comparably to wild-type (WT) Ft LVS. Collectively, these data indicate that the primary macrophage response to Ft LVS is overwhelmingly TLR2-dependent, requires de novo bacterial protein synthesis, and is independent of intracellular Ft replication.

Infect Immun