Internalization and phagosome escape required for Francisella to induce human monocyte IL-1β processing and release

doi:10.1073/pnas.0504271103

Internalization and phagosome escape required for Francisella to induce human monocyte IL-1β processing and release

^*Davis Heart and Lung Research Institute, ^‡Department of Molecular Virology and Immunology and Medical Genetics, and ^§Center for Microbial Interface Biology, Ohio State University, Columbus, OH 43210; and ^†Columbus Children's Research Institute, Columbus, OH 43205

Edited by Charles A. Dinarello, University of Colorado Health Sciences Center, Denver, CO, and approved November 15, 2005 (received for review May 23, 2005)

Abstract

Macrophage responses to Francisella infection have been characterized previously by subdued proinflammatory responses; however, these studies have generally focused on macrophage cell lines or monocyte-derived macrophages. Therefore, we studied the ability of fresh human blood monocytes to engulf and respond to Francisella by using the live vaccine strain variant and Francisella novicida. Because Francisella organisms have been reported to escape from the phagolysosome into the cytosol, we hypothesized that this escape may trigger the activation of caspase-1. Francisella tularensis variants were readily taken up by fresh human CD14⁺ monocytes, inducing the release of IL-1β, as well as IL-8, in a time- and dose-dependent fashion. Importantly, whereas live and dead Escherichia coli, F. novicida, and live vaccine strain, as well as the LPS of E. coli, were able to induce abundant IL-1β mRNA synthesis and intracellular pro-IL-1β production, only live Francisella induced enhanced IL-1β processing and release (51 ± 10 vs. 7.1 ± 2.1 ng/ml, for F. novicida vs. E. coli LPS; P = 0.0032). Cytochalasin D blocked the Francisella internalization and the Francisella-induced monocyte IL-1β processing and release but not that induced by the exogenous stimulus E. coli LPS. Also, killing bacteria did not block uptake but significantly diminished the IL-1β processing and release that was induced by Francisella. Blocking bacterial escape from the phagosome into the cytosol also decreased IL-1β but not IL-8 release. These findings demonstrate that Francisella organisms efficiently induce IL-1β processing and release in fresh monocytes by means of a sensing system that requires the uptake of live bacteria capable of phagosome escape.

Footnotes

↵ ¶ To whom correspondence should be addressed. E-mail: wewers.2{at}osu.edu.
Author contributions: M.A.G., I.J.B., and M.D.W. designed research; M.A.G., N.L.K., M.D.D., and M.W.H. performed research; J.S.G. contributed new reagents/analytic tools; and M.A.G. and M.D.W. analyzed data and wrote the paper.
Conflict of interest statement: No conflicts declared.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: moi, multiplicity of infection; LVS, live vaccine strain; RCN, relative copy number; PFM, paraformaldehyde; TLR, Toll-like receptor; NLR, NACHT leucine-rich repeats; cD, cytochalasin D.