Expression of Raf kinase inhibitor protein is downregulated in response to Newcastle disease virus infection to promote viral replication

Abstract

Newcastle disease virus (NDV) causes a severe and economically significant disease affecting almost the entire poultry industry worldwide. However, factors that affect NDV replication in host cells are poorly understood. Raf kinase inhibitory protein (RKIP) is a physiological inhibitor of c-RAF kinase and NF-kB signalling, known for their functions in the control of immune response as well as tumour invasion and metastasis. In the present study, we investigated the consequences of overexpression of host RKIP during viral infection. We demonstrate that NDV infection represses RKIP expression thereby promoting virus replication. Experimental upregulation of RKIP in turn acts as a potential antiviral defence mechanism in host cells that restricts NDV replication by repressing the activation of Raf/MEK/ERK and IkBa/NF-kB signalling pathways. Our results not only extend the concept of linking NDV-host interactions, but also reveal RKIP as a new class of protein-kinase-inhibitor protein that affects NDV replication with therapeutic potential.

Enhanced Replication of Virulent Newcastle Disease Virus in Chicken Macrophages Is due to Polarized Activation of Cells by Inhibition of TLR7

“…Consistent with previous studies (59,60), the amount of viral RNA in the cell and infectious virus titers in the supernatant significantly increased from 12 to 72 hpi in HD11 cells after infection with NDV strains (Figures 2A,B), although no infectious virus was detected in the supernatant at 4 hpi. While no differences in replication kinetics were observed between the two lentogenic NDV strains from 24 to 72 hpi, NDV strain LaSota presented a reduced ability to replicate in chicken macrophages (HD11) at 4 and 12 hpi, as evidenced by lower viral genomic RNA loads, when compared with the lentogenic NDV strain F55 (Figure 2A).…”

A Raf kinase inhibitor demonstrates antiviral activities both in vitro and in vivo against different genotypes of virulent Newcastle disease virus

“…To remove the cell debris, the homogenates were centrifuged for 15 minutes at 2,000 rpm, and the supernatant was collected for the virus titration. The virus titer was quantified by the micro-HA method as described previously (Yin et al, 2015). In line with the antiviral effects in vitro, treatment with a single dose of 10 mM RKIV resulted in a strong suppression for the virus replication in the lungs of the F48E9-, NA-1-, CC-1-, and HL-1-infected chickens at 24 hpi ( Figure 1B).…”

“…Complementing our earlier findings (Yin et al, 2015), we now addressed the question whether or not the Raf kinase inhibitor compound RKIV ( Figure 1A), which functions in the same way as RKIP by targeting the intracellular Raf kinase (Tsai et al, 2008), is also effective against the current enzootic virulent NDV isolates. As representative virulent NDV strains, we used the chicken origin classical genotype IX virulent F48E9 strain (F48E9), two genotype VII virulent strains goose/China/Jilin/NA-1/1999 (NA-1) and chicken/China/Jilin/HL-1/2014 (HL-1), and a genotype VI virulent strain pigeon/China/Jilin/CC-1/2014 (CC-1).…”

“…Our previous study demonstrated that targeted overexpression of host RKIP, which subsequently results in decreased Raf/MEK/ERK signaling and IBa/NF-B pathway activation, leads to reduced NDV production in cell culture (Yin et al, 2015). This unexpected dependence of NDV replication on the activity of RKIP offers the intriguing possibility to target this signaling pathway for an antiviral intervention.…”

“…The antiviral activity of RKIV against the panel of NDV strains (multiplicity of infection, 0.001) in vitro was assessed in a multicycle virus cell-based cytopathic effect (CPE) reduction assay with a colorimetric readout using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfphenyl)-2H-tetrazolium/phenazin e methosulfate (MTS/PMS) method as described previously (Sun et al, 2015). In brief, for the CPE reduction assay, log10 dilutions of RKIV (1,000-0.1 nM) were combined with 0.001 multiplicity of infection (MOI) of NDV and added into a monolayer of DF-1 or CEF cells in 96-well, white, flat-bottom cell cultural plates.…”