Two Domains of the V Protein of Virulent Canine Distemper Virus Selectively Inhibit STAT1 and STAT2 Nuclear Import

Abstract

Canine distemper virus (CDV) causes in dogs a severe systemic infection, with a high frequency of demyelinating encephalitis. Among the six genes transcribed by CDV, the P gene encodes the polymerase cofactor protein (P) as well as two additional nonstructural proteins, C and V; of these V was shown to act as a virulence factor. We investigated the molecular mechanisms by which the P gene products of the neurovirulent CDV A75/17 strain disrupt type I interferon ( Virulent canine distemper virus (CDV) causes a severe systemic infection in dogs that is characterized by high fever, diarrhea, and pneumonia. Large-scale immunosuppression is a hallmark of infection, and some virus strains additionally invade the central nervous system to cause chronic demyelinating encephalitis. The molecular mechanisms differentiating virulent from attenuated strains are poorly understood. However, the fact that dogs can be protected from infection with virulent CDV by vaccination with attenuated strains suggests that reliable induction of adaptive immunity is possible, provided that the critical early stage of infection is successfully mastered by the host. During the early stage of infection, host defense depends on the innate immune system, which is also responsible for generating signals that activate the adaptive immune response (27). The interferons of type I (IFN-I, e.g., IFN-␣/␤) are a critical element of the innate immune defense against viruses (13,36,41). Virtually all nucleated cells are capable of sensing viral infection by receptors such as Rig-I, MDA-5, or Toll-like receptor-3 (16). Activation of these receptors initiates a signal cascade that results in transcription, translation, and release from the cells of IFN-␣/␤. This part of the IFN defense is referred to as the induction stage. IFN action is initiated by the binding of IFN to type I IFN receptors that activates the receptor-associated tyrosine kinases JAK1 and Tyk2, which, in turn, phosphorylate the signal transducers and activators of transcription (STATs) (21, 41). Subsequently, the activated STAT1 and STAT2 together with IFN regulatory factor 9 (IRF9) form a complex, the IFN-stimulated gene factor 3 (ISGF3), which, once translocated to the nucleus, binds the IFN-stimulated response element (ISRE) sequence (39,45). This initiates the expression of well over 100 proteins which are responsible for the antiviral effect of IFN (36). In recent years, gene products targeting specific steps of IFN induction or action have been found in virtually all viruses studied, indicating the crucial role of IFN evasion in any successful interaction of viruses with their hosts.CDV, a Morbillivirus of the Paramyxoviridae, contains a nonsegmented, single-stranded, negative-sense RNA genome. The genome consists of six genes expressing the structural nucleocapsid (N), matrix (M), fusion (F), hemagglutinin (H), and large (L) proteins and the phospho-protein (P) (20). The P and the L proteins together form the RNA polymerase. In addition to the P protein, the nonstructural C and V p...

Morbillivirus Control of the Interferon Response: Relevance of STAT2 and mda5 but Not STAT1 for Canine Distemper Virus Virulence in Ferrets

“…Morbilliviruses interfere with IFN activation by preventing the nuclear translocation of STAT1 and STAT2 (28) as well as downstream signaling through the intracellular pattern recognition receptor mda5 (7,29). The V-protein residues involved in these functions have been mapped for MeV (8,19), and the importance of Y110H for STAT1 binding and the region for STAT2 and mda5 interactions has been demonstrated for other morbilliviruses (16,30). To investigate the roles of the different signaling pathways in morbillivirus pathogenesis, we selectively generated STAT1-, STAT2-, or mda5-blind CDV V proteins and a V protein with a disrupted zinc-binding domain (ZBDko) in the genetic background of the wild-type strain 5804P.…”

“…STAT1blind and STAT2blind CDV V proteins selectively lose their ability to prevent nuclear translocation of the respective STAT protein. Morbillivirus V proteins prevent type I and II IFN signaling by retaining STAT1 and STAT2 in the cytoplasm (16,28). To evaluate the extent of STAT1 and STAT2 nuclear translocation inhibition associated with the different mutants, the intracellular distribution of the respective proteins was quantified by confocal microscopy after stimulation with IFN-␥ and IFN-␣, respectively.…”

“…It is now clear that V protein inhibits the nuclear translocation of STAT1 and STAT2, thereby interfering with type I and II IFN-mediated transcriptional activation (16,28,30). In addition, V protein inhibits mda5 signaling (19).…”

“…The P/V shared domain alone can block type I and type II interferon (IFN) responses. This activity has been mapped to amino acids 110 to 130, with tyrosine 110 being essential for binding the signal transducer and activator of transcription 1 (STAT1) molecule (14)(15)(16). Inactivation of the zinc-binding domains in the V-protein unique region results in a 70% loss of its type I IFN inhibitory activity (8), and this region can directly inhibit beta interferon (IFN-␤) synthesis by interacting with the double-stranded RNA sensor melanoma differentiation-associated gene 5 (mda5) (7,17,18).…”

Paramyxovirus evasion of innate immunity: Diverse strategies for common targets

“…MeV V protein does not degrade STATs [104] , but has been reported by different laboratories to use several distinct mechanisms, including inhibition of STAT nuclear translocation without affecting STAT phosphorylation [103][104][105] , and inhibition of STAT1 and STAT2 phosphorylation due to interaction of its N-terminal domain with JAK1 [106,107] . Canine distemper virus (CDV) and RPV V proteins also inhibit IFN-activated STAT1/STAT2 nuclear import [108,109] , with RPV V protein, but not that of CDV, inhibiting STAT1/2 phosphorylation.…”