Activation of IκB Kinase by Herpes Simplex Virus Type 1

Abstract

Herpes simplex viruses (HSV) are ubiquitous pathogens causing a variety of diseases ranging from mild illness to severe life-threatening infections. HSV utilize cellular signaling pathways and transcription factors to promote their replication. Here we report that HSV type 1 (HSV-1) induces persistent activation of transcription factor NF-B, a critical regulator of genes involved in inflammation, by activating the IB kinase (IKK) in the early phase of infection. Activated NF-B enhances HSV-1 gene expression. HSV-1-induced NF-B activation is dependent on viral early protein synthesis and is not blocked by the anti-herpetic drug acyclovir. IKK inhibition by the anti-inflammatory cyclopentenone prostaglandin A 1 blocks HSV-1 gene expression and reduces virus yield by more than 3000-fold. The results identify IKK as a potential target for anti-herpetic drugs and suggest that cyclopentenone prostaglandins or their derivatives could be used in the treatment of HSV infection.One intriguing aspect of herpesviruses is their ability to influence host defense mechanisms and replication of other pathogens by inducing a stress response, via activation of cellular transcription factors, among which is nuclear factor-B (NF-B).1 NF-B is a critical regulator of the immediate-early pathogen response, playing an important role in promoting inflammation and viral gene expression (1). In most eukaryotic cells NF-B exists as an inactive cytoplasmic complex, whose predominant form is a heterodimer composed of p50 and p65 (Rel A) subunits, bound to inhibitory proteins of the IB family, usually IB␣ (1, 2). NF-B is activated in response to a variety of stress and pathogenic stimuli, including UV radiation, bacterial and viral infection, and proinflammatory cytokines (2, 3). Several stimuli activate NF-B by augmenting the activity of the IB kinase (IKK) complex, containing two catalytic subunits (IKK-␣ and IKK-␤) and the IKK-␥ or NEMO regulatory subunit (4, 5). IKK phosphorylates IBs at sites that trigger their ubiquitination and proteasome-mediated degradation (1, 4). Freed NF-B dimers translocate to the nucleus and activate a variety of genes encoding adhesion molecules, inflammatory and chemotactic cytokines, cytokine receptors, and enzymes that produce inflammatory mediators (1, 2). NF-B also activates the transcription of viral genes and is involved in several pathological events including progression of AIDS by enhancing human immunodeficiency virus type 1 (HIV-1) transcription (6, 7). Several viruses, including HIV-1 (8), cytomegalovirus (9), SV40 (10), and hepatitis B virus (11), contain functionally important NF-B-binding sites. In the case of HSV-1, NF-B-binding sites are located in the ICP0 and Vmw65 genes (12).In addition to containing NF-B-binding sites in its genome, HSV-1 can also induce NF-B nuclear translocation (13), and two immediate-early (IE) proteins, ICP4 and ICP27, were found to be required for this effect (13, 14); however, the signaling pathway utilized by the virus to activate the factor has still not been defin...

Protection by Herpes Simplex Virus Glycoprotein D against Fas-mediated Apoptosis

“…In the past, several reports have illustrated the capacity of HSV-1 to trigger NF-B activation following infection (42)(43)(44). A possible role for gD as a trigger in NF-B activation could be predicted, considering that HveA, which acts as a cell receptor for gD binding, activates NF-B through interaction with members of the tumor necrosis factor receptor-associated factor family (28) and following the engagement of its natural ligand, LIGHT (31).…”

“…Interestingly, it has been shown that HSV-1 infection activates NF-B (42,43). Particularly, it has been recently demonstrated that HSV-1 induces persistent activation of NF-B by activating the I B kinase in the early phase of infection of HEp-2 permissive cells (44). However, little information is available at the molec-ular level on mechanisms participating in this activation.…”

Herpes Simplex Virus Disrupts NF-κB Regulation by Blocking Its Recruitment on the IκBα Promoter and Directing the Factor on Viral Genes

“…Kinase Assay and Western Blot Analysis-Cell lysates were incubated with anti-IKK␣ antibodies in the presence of 15 l of protein-A-Sepharose at 4°C for 12 h. After washing, endogenous IKK activity was determined using GST-IB␣-(1-54) as substrate (11). For immunoblot analysis, equal amounts of protein (40 g/sample) from HaCaT whole cell extracts were separated by SDS-PAGE, blotted to nitrocellulose, and filters were incubated with polyclonal anti-IB␣/MAD3 (Santa Cruz Biotechnology), anti-IKK␣, or anti-p65 antibodies followed by decoration with peroxidase-labeled anti-mouse or anti-rabbit IgG (ECL, Amersham Biosciences) (11).…”

“…HaCaT cell monolayers were infected for 1 h at 37°C with HSV-1 strain F1 at a multiplicity of infection of 10 plaque forming units/ cell, unless stated otherwise. Virus titers were determined by plaque assay or by cytopathic effect 50% (CPE 50% ) assay on confluent VERO cell monolayers (11). Inactivated HSV-1 virus was prepared by exposure to UV light (254-nm wavelength) on ice for 15 min.…”

“…NF-B was found to be activated early during HSV-1 infection (10,11), and was shown to be involved in up-regulation of several host genes (12), as well as in promoting the progression of the virus replication cycle (11,13). However, the mechanisms governing NF-B activity in the nucleus of HSV-1-infected cells have still not been defined.…”