Bacterial secretion system skews the fate of Legionella-containing vacuoles towards LC3-associated phagocytosis.

The evolutionarily conserved processes of endosome-lysosome maturation and macroautophagy are established mechanisms that limit survival of intracellular bacteria. Similarly, another emerging mechanism is LC3-associated phagocytosis (LAP). Here we report that an intracellular vacuolar pathogen, Legionella dumoffii, is specifically targeted by LAP over classical endocytic maturation and macroautophagy pathways.

Autophagy activation by interferon-γ via the p38 mitogen-activated protein kinase signalling pathway is involved in macrophage bactericidal activity.

Macrophages are involved in many essential immune functions. Their role in cell-autonomous innate immunity is reinforced by interferon-γ (IFN-γ), which is mainly secreted by proliferating type 1 T helper cells and natural killer cells. Previously, we showed that IFN-γ activates autophagy via p38 mitogen-activated protein kinase (p38 MAPK), but the biological importance of this signalling pathway has not been clear.

Observation of autophagosome maturation in the interferon-γ-primed and lipopolysaccharide-activated macrophages using a tandem fluorescent tagged LC3.

Macrophages are engaged in many essential host functions, and their activation is a dynamic process that results in diverse functional outcomes such as the potentiation of bactericidal activity and production of chemokines, cytokines, and mediators that coordinate the inflammatory response. This pro-inflammatory response is bimodal, comprising a "prime" event, classically through interferon-γ (IFN-γ), and a "trigger," such as lipopolysaccharide (LPS). Recently, autophagy, which is one of the major degradative pathways in eukaryotic cells, has been shown to play an important role in both IFN-γ-primed and LPS-activated macrophages.

IFN-γ elicits macrophage autophagy via the p38 MAPK signaling pathway.

Autophagy is a major innate immune defense pathway in both plants and animals. In mammals, this cascade can be elicited by cytokines (IFN-γ) or pattern recognition receptors (TLRs and nucleotide-binding oligomerization domain-like receptors). Many signaling components in TLR- and nucleotide-binding oligomerization domain-like receptor-induced autophagy are now known; however, those involved in activating autophagy via IFN-γ remain to be elucidated.

Targeting of the GTPase Irgm1 to the phagosomal membrane via PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3) promotes immunity to mycobacteria.

Vertebrate immunity to infection enlists a newly identified family of 47-kilodalton immunity-related GTPases (IRGs). One IRG in particular, Irgm1, is essential for macrophage host defense against phagosomal pathogens, including Mycobacterium tuberculosis (Mtb). Here we show that Irgm1 targets the mycobacterial phagosome through lipid-mediated interactions with phosphatidylinositol-3,4-bisphosphate (PtdIns(3,4)P(2)) and PtdIns(3,4,5)P(3).

Emerging themes in IFN-gamma-induced macrophage immunity by the p47 and p65 GTPase families.

Vertebrates have evolved complex immune specificity repertoires beyond the primordial components found in lower multi-cellular organisms to combat microbial infections. The type II interferon (IFN-gamma) pathway represents one such system, bridging innate and acquired immunity and providing host protection in a cell-autonomous manner. Recent large-scale transcriptome analyses of IFN-gamma-dependent gene expression in effector cells such as macrophages have highlighted the prominence of two families of GTPases -- p47 IRGs and p65 GBPs -- that are now beginning to emerge as major determinants of antimicrobial resistance.

Enteropathogenic Escherichia coli, Shigella flexneri, and Listeria monocytogenes recruit a junctional protein, zonula occludens-1, to actin tails and pedestals.

Enteropathogenic Escherichia coli, Shigella flexneri, and Listeria monocytogenes induce localized actin polymerization at the cytoplasmic face of the plasma membrane or within the host cytoplasm, creating unique actin-rich structures termed pedestals or actin tails. The process is known to be mediated by the actin-related protein 2 and 3 (Arp2/3) complex, which in these cases acts downstream of neural Wiskott-Aldrich syndrome protein (N-WASP) or of a listerial functional homolog of WASP family proteins. Here, we show that zonula occludens-1 (ZO-1), a protein in the tight junctions of polarized epithelial cells, is recruited to actin tails and pedestals.

Assembly of the type III secretion apparatus of enteropathogenic Escherichia coli.

Enteropathogenic Escherichia coli (EPEC) secretes many Esps (E. coli-secreted proteins) and effectors via the type III secretion (TTS) system. We previously identified a novel needle complex (NC) composed of a basal body and a needle structure containing an expandable EspA sheath-like structure as a central part of the EPEC TTS apparatus.

BopC is a novel type III effector secreted by Bordetella bronchiseptica and has a critical role in type III-dependent necrotic cell death.

In Bordetella bronchiseptica, the functional type III secretion system (TTSS) is required for the induction of necrotic cell death in infected mammalian cells. To identify the factor(s) involved in necrotic cell death, type III-secreted proteins from B. bronchiseptica were analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and electrospray ionization tandem mass spectrometry.

Binding of intimin with Tir on the bacterial surface is prerequisite for the barrier disruption induced by enteropathogenic Escherichia coli.

Enteropathogenic Escherichia coli (EPEC) infects intestinal epithelial cells and perturbs the intestinal barrier that limits the paracellular movement of molecules. The disruption of the barrier is mediated by the effectors translocated into the host cells through the bacterial type III secretion system (TTSS). A previous report has described the importance of a bacterial outer membrane protein, intimin, in EPEC-mediated disruption of the barrier, and proposed that intimin, in concert with a host intimin receptor, controls the activity of the translocated barrier-disrupting effectors [P.

Enteropathogenic Escherichia coli type III effectors EspG and EspG2 alter epithelial paracellular permeability.

Enteropathogenic Escherichia coli (EPEC) delivers a subset of effectors into host cells via a type III secretion system. Here we show that the type III effector EspG and its homologue EspG2 alter epithelial paracellular permeability. When MDCK cells were infected with wild-type (WT) EPEC, RhoA was activated, and this event was dependent on the delivery of either EspG or EspG2 into host cells.

Type-III effectors: sophisticated bacterial virulence factors.

Bacterial pathogens cause a wide spectrum of diseases in human and other animals. Some virulence factors, which are referred to as effectors, are directly translocated into the host cell via an injection apparatus, i.e.

Enteropathogenic Escherichia coli activates the RhoA signaling pathway via the stimulation of GEF-H1.

Enteropathogenic Escherichia coli delivers a subset of effectors into host cells via a type III secretion system, and this step is required for the progression of disease. Here, we show that the type III effectors, EspG and its homolog Orf3, trigger actin stress fiber formation and the destruction of the microtubule networks beneath adherent bacteria. Both effectors were shown to possess the ability to interact with tubulins, and to stimulate microtubule destabilization in vitro.

The type III secreted protein BopD in Bordetella bronchiseptica is complexed with BopB for pore formation on the host plasma membrane.

The cytotoxicity of Bordetella bronchiseptica to infected cells is known to be dependent on a B. bronchiseptica type III secretion system. Although BopB, BopN, BopD, and Bsp22 have been identified as type III secreted proteins, these proteins remain to be characterized.

Bordetella dermonecrotic toxin undergoes proteolytic processing to be translocated from a dynamin-related endosome into the cytoplasm in an acidification-independent manner.

Bordetella pertussis dermonecrotic toxin (DNT), which activates intracellular Rho GTPases, is a single chain polypeptide composed of an N-terminal receptor-binding domain and a C-terminal enzymatic domain. We found that DNT was cleaved by furin, a mammalian endoprotease, on the C-terminal side of Arg(44), which generates an N-terminal fragment almost corresponding to the receptor-binding domain and a C-terminal remainder (deltaB) containing the enzymatic domain. These two fragments remained associated even after the cleavage and made a nicked form.

Identification of a receptor-binding domain of Bordetella dermonecrotic toxin.

Bordetella dermonecrotic toxin (DNT) stimulates the assembly of actin stress fibers and focal adhesions by deamidating or polyaminating Gln63 of the small GTPase Rho. DNT is an A-B toxin which is composed of an N-terminal receptor-binding (B) domain and a C-terminal enzymatically active (A) domain. In this study, to analyze the functional and structural organization of DNT, we prepared 10 clones of hybridoma producing anti-DNT monoclonal antibodies.

In vivo modifications of small GTPase Rac and Cdc42 by Bordetella dermonecrotic toxin.

Bordetella dermonecrotic toxin (DNT) is known to activate the small GTPase Rho through deamidation or polyamination. In this study, we examined whether Rac and Cdc42, the two other members of the Rho family, serve as intracellular targets for the toxin. Immunoprecipitation and immunoblot assays revealed that DNT deamidated or polyaminated intracellular Rac and Cdc42.