MxA-independent inhibition of Hantaan virus replication induced by type I and type II interferon in vitro

Interferons (IFN) induce an antiviral state against Hantaan virus (HTNV) but the mechanisms responsible for inhibition are unclear. The IFN-inducible MxA is discussed to be important for control of infections with hantaviruses. To characterize the role of endogenous MxA, the inhibition of HTNV induced by type I and type II IFNs was compared in Vero and A549 cells. IFNalpha and IFNgamma reduced production of infectious virions, viral RNA, and nucleocapsid protein with the same efficiency, although expression of MxA protein was detectable only in IFNalpha-treated A549 cells. Furthermore, knock down of MxA expression did not impair IFNalpha-induced inhibition. Thus, inhibition of HTNV induced by type I and type II IFNs did not dependent on expression of endogenous MxA. Taken together, these data suggest that MxA endogenously expressed in response to type I or type II IFNs does not play a pivotal role for the antiviral state against HTNV and that there is more than one mechanism by which cellular defences block hantavirus replication.     

Role for herpes simplex virus 1 ICP27 in the inhibition of type I interferon signaling

Host cells respond to viral infection by many mechanisms, including the production of type I interferons which act in a paracrine and autocrine manner to induce the expression of antiviral interferon-stimulated genes (ISGs). Viruses have evolved means to inhibit interferon signaling to avoid induction of the innate immune response. Herpes simplex virus 1 (HSV-1) has several mechanisms to inhibit type I interferon production, the activities of ISGs, and the interferon signaling pathway itself. We report that the inhibition of the Jak/STAT pathway by HSV-1 requires viral gene expression and that viral immediate-early protein ICP27 plays a role in downregulating STAT-1 phosphorylation and in preventing the accumulation of STAT-1 in the nucleus. We also show that expression of ICP27 by transfection causes an inhibition of IFN-induced STAT-1 nuclear accumulation. Therefore, ICP27 is necessary and sufficient for at least some of the effects of HSV infection on STAT-1.     

Inhibition by interferon of Sendai virus cytolysis.

Treatment of HeLa cells with human interferons inhibited 51Cr release from cells induced by ultraviolet-inactivated Sendai virus. The inhibitory effect became apparent about 6 h after interferon treatment and persisted for 24 to 48 h. In the interferon-treated cells, the cytolysis was inhibited within 10 min after adding virus and the inhibitory action was suppressed by the treatment of the cells with cycloheximide. Mock interferon and mouse interferon did not inhibit the cytolysis and antiinterferon serum neutralized the effect of interferon. All these findings indicate that Sendai virus-induced cytolysis is inhibited by interferon per se. However, interferon did not have any influence on Sendai virus hemolysis.     

Effects of exogenous interferon on L cells persistently infected with Sendai virus

Summary A carrier culture of L cells persistently infected with Sendai virus (steady state) designated as L-Sendai_ts cells was established with a temperature-sensitive strain of the virus. When interferon was added to culture fluids from the start of the cultures at permissive (35° C) or non-permissive temperature (38° C), cell-associated infectivity was unaffected at 35° C, while it was unexpectedly enhanced at 38° C, although the cell-associated infectivity was titrated after further incubation at 32° C for 2 days. The titer of cell-associated infectivity was increased by subculturing in the continuous presence of interferon at 38° C. The effect of interferon on the paradoxical enhancement of cell-associated infectivity was shown to be dose dependent. When L-Sendai_ts cells were successively subcultured 6 times at 38° C in the continuous presence or absence of interferon, more than 95 per cent of the cells contained a detectable amount of nucleocapsid (NP) antigen in the presence of interferon, whereas the antigen could be detected in only 30–40 per cent of the cells subcultured in the absence of interferon. Only when the cells subcultured at 38° C in the presence of interferon were transferred to permissive temperature, could the distinct hemadsorbing and cell-associated hemagglutinating activities and the release of virus particles, as measured by hemagglutinating activity in the culture fluids, be detected. Cells subcultured in the presence of interferon accumulated more virus polypeptides than in the absence of interferon. Accumulation of virus specific RNA in the cells subcultured in the presence of interferon was about twice as much as that in the absence of interferon. Larger sized RNA (probably 50S) was the major species and two smaller RNAs could be detected in both the treated and untreated cells.When L-Sendai_ts cells were cultured at 38° C in the presence of interferon, their multiplication was clearly inhibited. However, the cells which were subcultured twice at 38° C in the continouos presence of interferon acquired resistance to the anti-cell proliferative action of interferon. Interestingly, the conversion of the sensitive state to resistant state of the cells was reversible.With 3 Figures     

The C protein of measles virus inhibits the type I interferon response

Type I interferons (IFNalpha/beta) are an important part of innate immunity to viral infections because they induce an antiviral response and limit viral replication until the adaptive response clears the infection. Since the nonstructural proteins of several paramyxoviruses inhibit the IFNalpha/beta response, we chose to explore the role of the C protein of measles virus (MV) in such inhibition. Previous studies have suggested that the MV C protein may serve as a virulence factor, but its role in the pathogenesis of MV remains undefined. In the present study, a recombinant MV strain that does not express the C protein (MV C-) and its parental strain (Ed Tag) were used. Growth of MV C- was restricted in human peripheral blood mononuclear cells and HeLa cells, but in the presence of neutralizing antibodies to IFNalpha/beta, MV C- produced titers that were equivalent to those of Ed Tag. In addition, expression of the MV C protein from plasmid DNA inhibited the production of an IFNalpha/beta responsive reporter gene and, to a lesser extent, inhibited an IFNgamma responsive reporter gene. The ability of the MV C protein to suppress the IFNalpha/beta response was confirmed using a biologic assay. After IFNbeta stimulation, HeLa cells infected with Ed Tag produced five-fold less IFNalpha/beta than cells infected with MV C-. While the mechanism of inhibition remains unclear, these data suggest that the MV C protein plays an important role in the pathogenesis of MV by inhibiting IFNalpha/beta signaling.     

On the study of Sendai virus hemolysis. I. Complete Sendai virus lacking in hemolytic activity.

The hemolytic activity of Sendai virus was examined with samples collected at various times after infection of the embryonated eggs. The virus obtained just after the end of the one-step growth cycle (early harvest) was shown to exhibit no hemolytic activity, whereas it expressed full activity of cell fusion and infectivity. Incubation at 36°, freezing and thawing, or sonic treatment enabled the early harvests to be hemolytic. Conversion of the virus from the nonhemolytic state to a hemolytic one was always followed by the alteration of the virus envelope so that it became permeable to the uranyl acetate stain used for the negative staining (UA-particle). The degree of hemolysis and the number of UA-particles of the early harvest increased as freezing and thawing was repeated while the degree of cell fusion and infectivity was decreasing. On the basis of these findings, it was concluded that Sendai virus with intact envelope does not exhibit hemolysis, while it has the potential activity to do so.     

Enhanced endogenous type I interferon cell-driven survival and inhibition of spontaneous apoptosis by Riluzole

Highly active antiretroviral therapy (HAART), although effective in improving the survival of HIV-1-infected individuals, has not been able to reconstitute the adaptive immune response. We have described the use of novel chemical agents to restore T-cell survival/proliferation by inducing cytokine production. Due to its cationic amphiphilic structure, these molecules appear to enhance immune restoration. In this study, we investigated the action of Riluzole (2-amino-6-trifuromethoxybenzothiazole) in HIV-1 infection. Riluzole is able to increase (effective dose from 1 to 1000 nM) the cell-survival of T cells from HIV-1-infected patients and inhibit spontaneous apoptosis. The immunomodulatory effect of riluzole-sensitized cells was ascribed to endogenous type I interferon (IFN) derived from monocytes. Riluzole might be used for restoring the cell survival of immunocompromised patients and eliminating latent infected cells upon HIV-1 reactivation.     

Fusion inhibition: bioassay of type C viral protein.

Based on the observation that concentrated RD-114 virus directly induces fusion in a Rous sarcoma virus-transformed human glioma cell line, KC, an RD-114 virus protein was identified which had the property of inhibiting this fusion. The protein was irreversibly denatured by 2-mercaptoethanol, reacted with neutralizing sera to RD-114 virus, and eluted from guanidine agarose columns in a single peak in the 60 to 80,000 molecular-weight range. It does not prevent the fusion of KC cells by B-propiolactone-inactivated Sendai virus, and remained bound to KC cells even after extensive washing. It is suggested that the RD-114 fusion-inhibition factor is a viral surface protein which normally plays a role in virus attachment and is analogous to the 70,000-dalton glycoproteins (gp70) described for the murine Type C viruses.     

Wild-type rabies virus phosphoprotein is associated with viral sensitivity to type I interferon treatment

Rabies virus (RABV) is a single-stranded, negative-sense RNA virus that causes a fatal neurological disease in humans and animals. Our previous studies have shown that lab-adapted, but not wild-type (wt), RABV enhances innate immune responses including type I interferon (IFN) and chemokines. To determine if treatment with type I IFN can inhibit RABV infection, mouse neuroblastoma and baby hamster kidney cells were treated with IFN-α before being infected with lab-adapted or wt RABV. It was found that lab-adapted, but not the wt, RABV was able to replicate in IFN-α-pretreated cells. To determine the genes in wt RABV that confer sensitivity to IFN-α treatment, the P and the glycoprotein (G) genes from the wt RABV were used to replace the respective genes in the lab-adapted RABV. The results revealed that it is the P, not the G, gene that is associated with IFN sensitivity. Further studies have identified the regions containing the self-association domain (residues 59-139) and the C-terminal (residue 175-297) region on the P that might be associated with IFN sensitivity. The expression of ISGs, such as ISG15, ISG56, PKR, OAS-1G, was also investigated and found to be greatly increased in wt, but not in lab-adapted RABV-infected cells. It is possible that the P protein from the lab-adapted RABV can interfere with the downstream events in the interferon-signaling cascade.     

Effect of interferon on Vero cells persistently infected with Sendai virus compared to Vero cells persistently infected with SSPE virus

Summary Persistent infections with Sendai and SSPE virus were established in Vero cells. Sequential passages of these cells were monitored by immunofluorescence and for their sensitivity to the antiviral and antiproliferative effects of interferon (IFN). The cells rapidly developed resistance to the antiviral effect of IFN as judged by the inability of IFN to inhibit the replication of exogenous Sindbis virus. This decrease was accompanied by a reduction in the induction of the 2–5 oligo A synthetase. Both cell lines were resistant to the antiproliferative effect of IFN. A decrease or absence of IFN receptors on the surface of the cells was not found to be the cause of their resistance to IFN.     

Chicken interferon type I inhibits infectious bronchitis virus replication and associated respiratory illness.

Infectious bronchitis virus (IBV) causes an economically important respiratory disease in poultry worldwide. Previous studies have shown that CD8(+) cytotoxic T lymphocytes (CTL) are critical in controlling acute IBV infection, but the role of innate immunity is unknown. This study describes the in vitro and in vivo anti-IBV activity of natural spleen cell-derived and recombinant chicken interferon type I (rChIFN-alpha). Both natural and rChIFN-alpha inhibited replication of the Beaudette strain of IBV in chicken kidney cells (CKC) in a dose-dependent manner, with the antiviral activity of the former accounted for entirely by its content of type I IFN. IFN at 100 U/ml reduced viral replication by 50% as measured by syncytia formation. In addition, the spleen cell-derived supernatants (natural IFN) inhibited tracheal ring ciliostasis mediated by the Gray strain of IBV. Optimal protection against IBV-induced respiratory disease was obtained after intravenous or oral administration of ChIFN given 1 day before virus challenge and each of 5 days thereafter. ChIFN-I protected chicks from clinical illness by delaying the onset of the disease and decreasing the severity of illness, demonstrating its potential as an immune enhancer.     

Simple method for measuring growth inhibition by interferon of cells in monolayer

A method for determining cell growth inhibition by interferon based on a dye uptake and dilution technique is described. The results of this technique are comparable to cell counting and measurement of 3H-TdR incorporation, but the technique is easier and more rapid to perform.