Interferon-beta-mediated Innate Host Defense in Rickettsia conorii-infected Human Endothelium

Rickettsia conorii, an obligate intracellular bacterium and the causative agent of Mediterranean spotted fever, preferentially infects the human microvascular endothelium to activate proinflammatory and innate immune responses, as evidenced by enhanced expression and secretion of cytokines and chemokines upon infection. Our aim was to determine whether Interferon (IFN)-β, a cytokine traditionally considered to be involved in antiviral immunity, would establish a cell-intrinsic, innate host defense against R. conorii in infected human endothelial cells. Results indicate that IFN-β secretion does induce STAT1 protein phosphorylation at Tyrosine 701 and Serine 727 via the JAK-STAT signaling pathway. Employing siRNA-directed gene knock-down techniques, recombinant IFN-β treatments, and IFN-β neutralizing antibodies, we confirmed that IFN-β-mediated STAT1 activation has a protective role against intracellular R. conorii replication. The presence of viable intracellular rickettsiae was essential for the induction of innate immune responses; inhibition of rickettsial metabolic activity by tetracycline treatment inhibited both IFN-β expression and STAT1 phosphorylation.;The infected endothelium induces the expression of IFN-stimulated genes in an IFN β-dependent manner, including ISG15, UBP43, OAS1, MX1, IRF1, IRF9, and TAP1. Increased expression of both free ISG15 and conjugated-ISG15, targeting yet unidentified proteins, was observed in cultured human endothelial cells. UBP43, an ISG15-specific deconjugating enzyme, maintained the balance between free and conjugated ISG15 in infected cells. ISG15 knock-down by siRNA transfection resulted in a significant increase in the extent of rickettsial replication, whereas UBP43 knock-down yielded a reciprocal inhibitory effect, indicating that both ISG15 and UBP43 play a role in antirickettsial immunity. In addition to ISG15 deconjugation, UBP43 also functions as a negative regulator for IFN-β pathway. Confirming its negative regulatory role, UBP43 knock-down led to increased STAT1 phosphorylation and transcriptional activation of IFN-stimulated genes such as OAS1, MX1, and GBP1. Expression of SOCS1, a known negative regulator for the JAK-STAT signaling pathway, was also induced by IFN-β-dependent and independent mechanisms. However, STAT1 activation was not affected by SOCS1 knock-down in infected cells. In summary, we concluded that induction of specific IFN-stimulated genes by IFN-β secretion inhibits R. conorii replication, providing a new insight into the host-pathogen interactions of the spotted fever group rickettsiae.