Lipidation of the FPI protein IglE contributes toFrancisella tularensisssp.novicidaintramacrophage replication and virulence

Abstract

Francisella tularensis is a Gram-negative bacterium responsible for the human disease tularemia. The Francisella pathogenicity island (FPI) encodes a secretion system related to type VI secretion systems (T6SS) which allows F. tularensis to escape the phagosome and replicate within the cytosol of infected macrophages and ultimately cause disease. A lipoprotein is typically found encoded within T6SS gene clusters and is believed to anchor portions of the secretion apparatus to the outer membrane. We show that the FPI protein IglE is a lipoprotein that incorporates 3H-palmitate and localizes to the outer membrane. A C22G IglE mutant failed to be lipidated and failed to localize to the outer membrane, consistent with C22 being the site of lipidation. Francisella tularensis ssp. novicida expressing IglE C22G is defective for replication in macrophages and unable to cause disease in mice. Bacterial two-hybrid analysis demonstrated that IglE interacts with the C-terminal portion of the FPI inner membrane protein PdpB, and PhoA fusion analysis indicated the PdpB C-terminus is located within the periplasm. We predict this interaction facilitates channel formation to allow secretion through this system.

Francisella tularensistype VI secretion system comes of age

“…Deletion of iglE genes blocks intracellular proliferation of virulent F. tularensis strain as well as leads to in vivo strain attenuation in mice. 26 The defect in intracellular replication and virulence is associated with the loss of IglE lipidation that is responsible for its localization to bacterial outer membrane. Two-hybrid and in vivo co-immunoprecipitation analyses documented that membrane bounded IglE interacts with C-terminus of PdpB/Icm and by this way contributes to channel formation and protein secretion through the Francisella T6SS.…”

“…The previously published data on both the F. tularensis Schu S4 and the F. novicida IglE mutant described the defective bacterial replication in bone-derived macrophages and within J774 cells, respectively. 26,27 Br€ oms et al study exploiting direct injection of the IglE mutant into the cytoplasm of host cells to avoid of phagosome compartment proved efficient cytosolic replication of the mutant strain. Hence, it is evident that IglE protein is important for the phagosomal escape but not for subsequent cytosolic replication.…”

Enhancement of vaccine efficacy by expression of a TLR5 ligand in the defined live attenuated Francisella tularensis subsp. novicida strain U112ΔiglB::fljB

“…Constitutive expression of the FljB D1 domain in the generated U112▲iglB::fljB was under the control of the Francisella heat-shock protein groE gene promoter. To confirm the expression of FljB in U112▲iglB::fljB, we used anti-FljB antibody to detect the protein in the bacterial cytosolic and membrane fractions prepared according to our previous publication [20]. As shown in Fig.…”

Expression of Francisella pathogenicity island protein intracellular growth locus E (IglE) in mammalian cells is involved in intracellular trafficking, possibly through microtubule organizing center

“…Several reports indicate that IglE is a bacterial lipoprotein with a signal peptide and is located at the bacterial membrane where it forms part of the T6SS apparatus (Bröms, Meyer, & Sjöstedt, ; Nguyen, Gilley, Zogaj, Rodriguez, & Klose, ; Robertson, Child, Ingle, Celli, & Norgard, ). In addition, we observed limited secretion of IglE into the culture medium.…”