Inhibition of IκB Kinase by Vaccinia Virus Virulence Factor B14

Abstract

The IκB kinase (IKK) complex is a key regulator of signal transduction pathways leading to the induction of NF-κB-dependent gene expression and production of pro-inflammatory cytokines. It therefore represents a major target for the development of anti-inflammatory therapeutic drugs and may be targeted by pathogens seeking to diminish the host response to infection. Previously, the vaccinia virus (VACV) strain Western Reserve B14 protein was characterised as an intracellular virulence factor that alters the inflammatory response to infection by an unknown mechanism. Here we demonstrate that ectopic expression of B14 inhibited NF-κB activation in response to TNFα, IL-1β, poly(I:C), and PMA. In cells infected with VACV lacking gene B14R (vΔB14) there was a higher level of phosphorylated IκBα but a similar level of IκBα compared to cells infected with control viruses expressing B14, suggesting B14 affects IKK activity. Direct evidence for this was obtained by showing that B14 co-purified and co-precipitated with the endogenous IKK complex from human and mouse cells and inhibited IKK complex enzymatic activity. Notably, the interaction between B14 and the IKK complex required IKKβ but not IKKα, suggesting the interaction occurs via IKKβ. B14 inhibited NF-κB activation induced by overexpression of IKKα, IKKβ, and a constitutively active mutant of IKKα, S176/180E, but did not inhibit a comparable mutant of IKKβ, S177/181E. This suggested that phosphorylation of these serine residues in the activation loop of IKKβ is targeted by B14, and this was confirmed using Ab specific for phospho-IKKβ.

Curcumin Inhibits Rift Valley Fever Virus Replication in Human Cells

“…Interestingly, activated, phosphorylated IKK-␤ promotes viral replication, because the viral protein RTA (replication and transcription activator) is a phosphorylation target for IKK-␤ kinase activity (49). In contrast to this situation, in the case of vaccinia virus infection, the viral protein B14 modulates IKK-␤ kinase activity in a way that inhibits the phosphorylation of its downstream target IB␣ without exerting any influence on IKK-␣ activity (50,51). In a similar manner, enterovirus 71 2C protein is known to target IKK-␤ activity by interacting with IKK-␤, which is a critical event in the infectious process (52).…”

NFκB activation by modified vaccinia virus as a novel strategy to enhance neutrophil migration and HIV-specific T-cell responses

“…The A52, K7, and B15 early proteins, members of the B-cell lymphoma-2 family (31,32), are involved in suppression of host immune responses (2). During infection, A52 and K7 interact with TNF receptor-associated factor 6 and inhibit its kinase activation capacity (24,33), whereas B15 binds the IκB kinase complex to inhibit IκBα phosphorylation and degradation (20). In this study, we focused on the A52, K7, and B15 proteins; using single, double, and triple deletion mutants, we demonstrate that the presence of only one of the three inhibitory molecules is sufficient to abolish NFκB activation, ruling out synergy between these proteins as a mode of action.…”

“…VACV interacts with TLR2 (16) and TLR4 (17) and produces early direct or indirect viral inhibitors of the NFκB pathway such as A46 (18), A49 (19), A52 (18), B14 (20), C4 (21), E3 (22), K1 (23), K7 (24), M2 (25), and N1 (26). New York vaccinia virus (NYVAC), a highly attenuated VACV strain used as vaccine vector, lacks most of these proteins but encodes NFκB pathway Significance Although poxvirus vectors are widely used in preclinical and clinical trials as candidate vaccines for multiple pathogens, how these vectors affect the host immune response is not clear.…”

Poxviral Protein A46 Antagonizes Toll-like Receptor 4 Signaling by Targeting BB Loop Motifs in Toll-IL-1 Receptor Adaptor Proteins to Disrupt Receptor:Adaptor Interactions

“…VACV encodes immunomodulatory proteins which act intracellularly to target various components of innate immune signal transduction pathways. These viral proteins include A46 (32), A52 (33), N1 (34), B14 (35), K7 (36), and C6 (37). Of these, A46 and A52 specifically disrupt TLR signaling, and we previously demonstrated a role for A46 in VACV virulence (32).…”