The Immune Escape Strategy of Rabies Virus and Its Pathogenicity Mechanisms

Abstract

In contrast to most other rhabdoviruses, which spread by insect vectors, the rabies virus (RABV) is a very unusual member of the Rhabdoviridae family, since it has evolved to be fully adapted to warm-blooded hosts and spread directly between them. There are differences in the immune responses to laboratory-attenuated RABV and wild-type rabies virus infections. Various investigations showed that whilst laboratory-attenuated RABV elicits an innate immune response, wild-type RABV evades detection. Pathogenic RABV infection bypasses immune response by antagonizing interferon induction, which prevents downstream signal activation and impairs antiviral proteins and inflammatory cytokines production that could eliminate the virus. On the contrary, non-pathogenic RABV infection leads to immune activation and suppresses the disease. Apart from that, through recruiting leukocytes into the central nervous system (CNS) and enhancing the blood-brain barrier (BBB) permeability, which are vital factors for viral clearance and protection, cytokines/chemokines released during RABV infection play a critical role in suppressing the disease. Furthermore, early apoptosis of neural cells limit replication and spread of avirulent RABV infection, but street RABV strains infection cause delayed apoptosis that help them spread further to healthy cells and circumvent early immune exposure. Similarly, a cellular regulation mechanism called autophagy eliminates unused or damaged cytoplasmic materials and destroy microbes by delivering them to the lysosomes as part of a nonspecific immune defense mechanism. Infection with laboratory fixed RABV strains lead to complete autophagy and the viruses are eliminated. But incomplete autophagy during pathogenic RABV infection failed to destroy the viruses and might aid the virus in dodging detection by antigen-presenting cells, which could otherwise elicit adaptive immune activation. Pathogenic RABV P and M proteins, as well as high concentration of nitric oxide, which is produced during rabies virus infection, inhibits activities of mitochondrial proteins, which triggers the generation of reactive oxygen species, resulting in oxidative stress, contributing to mitochondrial malfunction and, finally, neuron process degeneration.

Keywords: BBB permeability; IFN-α/β; INOS; RABV; apoptosis; autophagy; cytkines/chemokines; immune evasion; mitochondrial dysfunction; neurons.

Figure 3

The method by which neural production of CXCL10 caused by laboratory-attenuated rabies virus (RABV) enhances the blood–brain barrier (BBB) permeability. 1. Chemokine CXCL10 is secreted by laboratory-attenuated RABV infected neurons; 2. CXCL10 mediates CD4⁺ T cell recruitment into the CNS; 3. CXCL10 mediates CD4⁺ T cell differentiation into Th1 and Th17 cells; 4a, IFN-γ secreting Th1 cells could further boost the induction of CXCL10 through positive feedback; 4b, IL-17 secreting Th17 cells alters the TJ (tight junction) proteins leading to Breakdown of the blood–brain barrier [67].

Figure 1

(A). RABV virion. Rabies virus is enveloped, bullet-shaped, with a body size of about 180 × 75 nm. (B). RABV genome. The RNA genome of the RABV is single-stranded and non-segmented, with a genome size of about 12 kb. It contains a leader and trailer region at the 3′ and 5′ ends, along with five structural proteins (N, P, M, G, and L) and four intergenic non-coding sequences. Multiple P proteins (P (P1), P2, P3, P4 and P5) are produced through alternative initiation from in-frame AUG start codons due to a leaky scanning mechanism. (Original source of the image; Philippe Le Mercier, SIB Swiss Institute of Bioinformatics) [9].

Figure 2

The various interactions between RABV proteins and interferon signaling pathway. The IFN-signaling pathway is activated by RABV infection, whereas RABV utilizes its own proteins to suppress the IFN signaling pathway by interacting with key factors in IFN pathways. RABV P interacts directly with tyrosine-phosphorylated STAT (pY-STAT) to inhibit the expression of ISGs (interferon-stimulated genes) by affecting its localization and reducing its ability to bind to ISG promoters. P protein is also able to interact directly with TBK-1 (TANK-binding kinase-1) in a dose-dependent manner, inhibiting phosphorylation of the IRF3 (interferon regulatory factor 3). Moreover, the interaction between P and PML (promyelocytic leukemia) alters PML protein localization and the structure of PML-NBs (nuclear bodies), thereby regulating IFN-induced apoptosis. The encapsulation of RABV RNA by N protein protects viral RNA from getting recognized by RIG-I (retinoic acid–inducible gene I), which subsequently prevents RIG-I-mediated activation of the downstream IRF-3 pathway. Type-I IFN stimulation causes the M to shift toward activated pSTAT1 interaction, which improves the ability of P protein to engage with JAK1 to prevent pSTAT1 from activating and with pSTAT1 to restrain it in the cytoplasm [43].

Figure 4

Mechanisms of mitochondrial dysfunction associated with rabies virus infection. In mitochondria of rabies virus-infected neurons, the activity of electron transport system (ETS) complexes I and IV are increased either due to direct (e.g., interaction with a rabies virus protein) and/or indirect (e.g., high NADH/NAD⁺ ratio and up-regulation of sirtuin activity) viral effect. The increased proton pumping across mitochondrial membranes generates a high mitochondrial membrane potential (MMP). Electron leakage during both forward and reverse ETS leads to superoxide (O₂⁻) formation that is dismutated by mitochondrial superoxide dismutase (SOD₂) to hydrogen peroxide (H₂O₂). Both O₂⁻ and H₂O₂ may lower the intracellular ATP levels by increasing hydrolysis of ATP molecules. Overproduction and accumulation of O₂⁻ and H₂O₂ induces oxidative stress and leads to degeneration of neuronal processes (solid lines findings, dashed lines hypothesis) [110].

Figure 5

Intramitochondrial metabolism of nitric oxide and superoxide radicals. The steady-state concentrations are indicated. Oxygen and nitrogen-reactive species are kept in biological systems at steady-state concentrations that can be estimated by using the steady-state approach with the assumption that the rate of production is equal to the rate of utilization. The primary production of O₂⁻, cytosolic Cu-Zn-SOD and mitochondrial Mn-SOD keep steady-state concentrations of 10⁻¹⁰ M in the mitochondrial matrix and 10⁻¹¹ M in the cytosol. The cytosolic steady-state concentration of H₂O₂ estimated from the rate of H₂O₂ generation by subcellular sources and its removal by catalase and glutathione peroxidase is about 10⁻⁷–10⁻⁸ M. considering the rate of H₂O₂ production, its removal by intra-mitochondrial glutathione peroxidase and its diffusion to the cytosolic space, H₂O₂ steady-state concentration in the mitochondrial matrix results in approximately 10⁻⁸ M. The balance between NO production and its utilization by the reactions with the components of the respiratory chain and with O₂⁻ regulates the intra-mitochondrial steady-state concentration of NO at about 50 nM, which in turn regulates mitochondrial oxygen uptake and energy supply [137].

Figure 6

Modes of mitochondrial operation that lead to O₂^{• −} production. There are three modes of mitochondrial operation that are associated with O₂^{• −} production. In mode 1, the NADH pool is reduced, for example by a damage to the respiratory chain, loss of cytochrome c during apoptosis or low ATP demand. This leads to a rate of O₂^{• −} formation at the FMN of complex I that is determined by the extent of FMN reduction which is in turn set by the NADH/NAD⁺ ratio. In mode 2, there is no ATP production and there is a high proton motive force and a reduced CoQ (Ubiquinone) pool which leads to reverse electron transport (RET) through complex I, producing large amounts of O₂^{• −}. In mode 3, mitochondria are actively making ATP and consequently have a lower change in membrane proton motive (MPM) force than in mode 2 and a more oxidized NADH pool than in mode 1. Under these conditions, the flux of O₂^{• −} within mitochondria is far lower than in modes 1 and 2, and the O₂^{• −} sources are unclear [143].