Interferon-γ Listericidal Action Is Mediated by Novel Rab5a Functions at the Phagosomal Environment

Abstract

Control and clearance ofListeria monocytogenes is an intracellular facultative bacterium able to invade phagocytic cells and is responsible for severe pathologies in immunocompromised people, newborns and pregnant women (1). L. monocytogenes entry into the host cell is an active process involving several protein components. After a short phagosomal period (ϳ30 min), L. monocytogenes escapes to the cytosol, avoids intracellular killing, and replicates (reviewed in Ref. 2). The L. monocytogenes survival mechanism involves two steps: (i) live bacteria avoid phagosome maturation by inactivation of the endosomal trafficking regulator Rab5a, which blocks the recruitment of lysosomal proteins to the phagosomes (Lamp-1 and cathepsin-D) (3) and (ii) secretion by L. monocytogenes of listeriolysin and PI-PLC lyses the phagosomal membrane, translocates L. monocytogenes to the cytoplasm, and consequently, allows for L. monocytogenes intracellular survival (4).Control of L. monocytogenes infection and clearance is an interferon-␥ (IFN-␥) 1 -dependent process. IFN-␥ priming of macrophages (MØs) recruited at the inflammatory site triggers their listericidal abilities (5). IFN-␥ signaling modulates the expression and activation of more than 200 proteins (6). However, to date, only a few of these molecules have been shown to exert a direct role in pathogen elimination (7). Among these are (i) IGTP, a GTP-binding protein relevant for Toxoplasma clearance (8) and (ii) Nramp1, a MØ-restricted lysosomal protein involved in Leishmania, Salmonella, and Mycobacterium spp. clearance (9). In addition, IFN-␥ induces the production of reactive oxygen (ROI) and nitrogen (RNI) intermediates with microbicidal activity (10). From this set of molecules, only ROI and RNI have been shown to restrict L. monocytogenes growth (10, 11), while the other two molecules (i.e. IGTP or Nramp1) play no role at all in L. monocytogenes clearance (8, 9).Recently, we have shown that in resting MØs the inhibition of Rab5a synthesis allows for intracellular survival of a listeriolysin-defective L. monocytogenes mutant, that under normal Rab5a levels is unable to grow and fails to escape from the phagosome (12). Furthermore, we have also described that IFN-␥ signaling up-regulates Rab5a function (13). However, at this stage, no correlation between the induction of ROI and RNI by IFN-␥ and the Rab5a function has been established. Here, we show that Rab5a is a key molecule for the IFN-␥ promoted clearance of a pathogenic L. monocytogenes strain at the phagosomal stage. We show that Rab5a, in the GTP form, controls the recruitment of active Rac2 to the transformed L. monocytogenes phagolysosome and the assembly of the phagocyte NADPH oxidase with the production of toxic radicals. These Rab5a-mediated actions compromise Listeria viability within the phagolysosomes and further L. monocytogenes intracellular survival. EXPERIMENTAL PROCEDURESCells and Reagents-J774 cells and proteose peptone-elicited peritoneal MØs from Balb/c mice were cultured in Dulbecco's modified Eag...

IIGP, a member of the IFN inducible and microbial defense mediating 47 kDa GTPase family, interacts with the microtubule binding protein hook3

“…These IFN-induced mechanisms are in part reflected by profound changes in the dynamics and functional properties of distinct intracellular vesicles (Sidhu et al, 1999;AlvarezDominguez and Stahl, 1998;Tsang et al, 2000;Shurety et al, 2000). For example, the IFNγ-regulated GTPase Rab5a causes remodeling of the phagosomal environment, resulting in listeriocidal activity (Prada-Delgado et al, 2001). In analogy, IIGP could modulate processes of vesicular trafficking in which mHk3 is involved.…”

The IFN-Inducible Golgi- and Endoplasmic Reticulum- Associated 47-kDa GTPase IIGP Is Transiently Expressed During Listeriosis

“…It is tempting to speculate that these 47-kDa GTPases participate in these processes. In analogy, a member of the GTP-binding Rab family, Rab5a, localizes to early endosomes and has been shown to be involved in listericidal activity in macrophages (50).…”

Critical Role for Transcription Factor C/EBP-β in Regulating the Expression of Death-Associated Protein Kinase 1

“…In most cases, the critical transcription factor mediating the IFN effects had not been identified. Some of these gene products, such as RAC-2 and p130 (Rb related), are modulated by type I IFNs (60,87). The p8 protein codes for a homologue of a bovine chemokine (85).…”