Internalin Pathway
Explore and order pathway-specific siRNAs, real-time PCR assays, and expression vectors. View pathway information and literature references for your pathway.
  • Click on your proteins of interest in the pathway image or review below
  • Select your genes of interest and click "add selection"
  • When you have finished your gene selection, click "Find Products" to find assays, arrays, or create custom products
Download Image Terms of Use Download PPT
Pathway Navigator
Internalin Pathway

An essential step in the life cycle of many important pathogenic bacteria is their ability to invade cells that are normally nonphagocytic. Gaining access to an intracellular niche provides bacteria with an environment permissive for growth, allows them to avoid host defense mechanisms, or permits them to gain access to deeper tissues. Listeria monocytogenes is a food-borne intracellular pathogen that, although most often not life-threatening, can occasionally cause serious listeria infections in immunosuppressed individuals, pregnant women, and neonates (Ref.1). Critical for the development of a systemic infection is the ability of this bacterium to be internalized by both professional phagocytes and nonphagocytic cells. It is the ability of these bacteria to invade non-phagocytic cells, plays an essential role in breaching the natural barriers: the intestinal epithelium and the placental and blood-brain barriers (Ref.2). L. monocytogenes induces its own uptake into non-phagocytic host cells through the actions of InlA (Internalin-A) and InlB (Internalin-B), two related LRR (Leucine-Rich Repeat)-containing virulence factors that localize to the bacterial surface (Ref.3). These molecules mimic natural host cell ligands and trigger engulfment of the bacterium after specifically interacting with cell-surface receptors.

InlA-mediated entry is restricted to a few epithelial cells, whereas, InlB promotes entry into various cell types, such as hepatocytes, epithelial cells and endothelial cells. InlA is a ligand for E-cadherin, a cell adhesion molecule present in epithelial tissues and involved in the formation of intercellular junctions (Ref.4) InlB triggers bacterial entry by interacting with the HGFR (Hepatocyte Growth Factor Receptor)/Met and two other cellular components: gC1qR and proteoglycans/ GAGs (Glycosaminoglycans). gC1qR is a ubiquitous glycoprotein, that acts as a coreceptor for Met. Signaling events elicited by InlA or InlB lead to actin-mediated zippering of the host membrane around the bacterium and internalization. These induced phagocytosis processes require tightly regulated steps of actin polymerization and depolymerization, tyrosine phosphorylation, and PI3K (Phosphatidylinositiol-3-Kinase) activity (Ref.5). InlA and InlB share significant amino acid sequence similarity and the most important feature is the presence at their amino terminus of a domain composed of a varying number of LRRs. This domain is critical for the stimulation of bacterial entry. It is the composition and number of the LRRs that establishes the receptor specificity of these bacterial ligands, which, in turn, determines their individual contribution to bacterial entry into different mammalian cells (Ref.2). E-cadherin is connected to the actin cytoskeleton via its cytoplasmic carboxy-terminal domain and at least three cytoplasmic proteins: Alpha Catenin, Beta Catenin and Gamma Catenin. Althogh it is known that InlA-E-cadherin interaction, via Rac activation, can trigger dynamic events of actin polymerization and membrane extension, culminating in bacterial uptake, our knowledge of the exact procedure of signal transduction downstream of the InlA-E-cadherin interaction is still fragmentary (Ref.4). By contrast, signaling pathways activated by InlB have been dissected in more detail, revealing the strikingly potent signaling properties of InlB (Ref.3).

InlB, the first-identified bacterial agonist of a receptor tyrosine kinase is composed of an amino-terminal receptor-binding domain followed by a B repeat region (B) and three GW domains. The receptor-binding domain contains three motifs: an amino-terminal cap, a LRR segment, and an IR (Immunoglobin-like Region). While the amino-terminal cap and the LRR domain are sufficient to bind and activate the Met, the GW domains attach InlB non-covalently to the bacterial cell wall (Ref.6). In bacteria present in the environment, InlB is buried into the bacterial cell wall, somehow protected from proteolytic degradation and external aggressive agents. Upon infection, in proximity with the targeted host cell, by interacting with GAGs and gC1qR it becomes accessible to the Met receptor (Ref.3). InlB, when present at the bacterial surface, mediates entry into the host cell by zipper-type phagocytosis and soluble InlB induces membrane ruffling.  Both require actin polymerization, tyrosine phosphorylation and PI3K activation. A pool of the protein may also be released in the medium and induce signals as a prelude to or independently from entry (Ref.2).

InlB stimulates the sequential tyrosine phosphorylation of Met, recruitment and phosphorylation of several signaling molecules including GRB2 (Growth Factor Receptor-Bound Protein-2), GAB1 (GRB2 Associated Binding protein-1), SHC (SH2 Containing Protein), Crk, CrkL, Cbl, Rho-GTPases (Rac and CDC42) and activation of the p85-p110 PI3K. Activation of PI3K affects cytoskeletal dynamics and cell survival. GRB2 works together with SOS to link activated receptor protein-tyrosine kinases to Ras activation, followed by the activation of the MAPK (Mitogen-Activated Protein Kinase) cascade. Crk and CrkL are related adaptor proteins that are also involved in regulating cytoskeletal changes. Cbl is an ubiquitin ligase that regulates receptor internalization and degradation (Ref.7). PI3K plays multiple roles in InlB-mediated internalization, including recruitment of both membrane vesicles and actin regulatory proteins. Proteins of the Vav family, which act as GEFs (Guanine Exchange Facors) for Rac and are downstream effectors of PI3K signaling, have a role in Rac activation downstream of Met. Rac activation elicit a Rac-PAK(p21-Activated Kinase)-LIMK (LIM Kinase)-Cofilin cascade. In addition, ARP2/3 (Actin-Related Protein) complex regulates actin dynamics during InlB-induced phagocytosis. Activated Rac recruits IRSP53, which binds to the proline-rich region of WAVE (WASP Family Verprolin Homology Domain-Containing Protein), a member of the WASP (Wiskott-Aldrich Syndrome Protein) family. As a result, the C-terminal region of WAVE is exposed and activates the ARP2/3 complex. The ARP2/3 complex is an assembly of seven proteins that together promote nucleation of actin filaments and therefore participate in the formation of branched actin networks. All these events lead to the initiation of actin polymerization and actin rearrangements at the phagocytic cup leading to InlB-induced listeria uptake (Ref.1).

Actin polymerization is thought to provide the driving force that propels membranes around the bacterium. However, the shaping of the phagocytic cup also requires actin depolymerization events, particularly beneath the particle, to facilitate retraction of the cup. Proteins of the ADF (Actin Depolymerising Factor)/Cofilin family, are mediators of this process. These proteins increase the rate of actin turnover and the number of free actin ends available for polymerization. They are inactivated through LIMK-induced phosphorylation and reactivated through dephosphorylation. At low activity, controlled by LIMK, Cofilin could be involved in the phagocytic cup extension by stimulating actin dynamics. Then, dephosphorylation of Cofilin and its progressive accumulation on filaments would ultimately favor the disassembly of the actin network during the retraction of the phagocytic cup and around the newly formed phagosome (Ref.5). PI3K also functions in pseudopod extension and closure of the phagocytic cup by regulating exocytosis of endomembranes and membrane fusion events.

InlB, not only promotes the phagocytic events, but, as a soluble factor, it also activates signaling pathways that are not directly linked to phagocytosis. InlB-mediated Akt activation plays a role in the survival of the infected host cell. PI3K activates the Akt/ PKB (Protein Kinase-B), which activates the IKKs (I-KappaB Kinases) complex, followed by phosphorylation of IKappaB and the subsequent activation of the NF-KappaB (Nuclear Factor-KappaB) pathway. Since L. monocytogenes is an intracellular pathogen, host cell survival is important as it could facilitate the dissemination of the bacteria in tissues. Activation of PLC-Gamma1 (Phosholipase-C-Gamma1) results from its PI3K dependent association with tyrosine-phosphorylated proteins. InlB-induced PLC-Gamma1 activation and calcium mobilization are probably involved in post-internalization steps, such as the control of cell growth and/or of gene expression (Ref.3). Human listeriosis is characterized by a high mortality rate, with clinical features including meningitis or meningo-encephalitis, septicemia, in some cases gastroenteriris, abortions, and perinatal infections. InlA and InlB molecules pave the way to these serious listeria infections, by triggering the engulfment of the bacterium into non-phagocytic cells. A detailed study of this versatile family of bacterial proteins promises to uncover novel therapeutic avenues to combat listeriosis (Ref.6).