The Human RVB Complex Is Required for Efficient Transcription of Type I Interferon-Stimulated Genes

Abstract

Type I interferons (IFNs) stimulate transcription through a latent heterotrimeric transcription factor composed of tyrosinephosphorylated STAT1 and STAT2 and the DNA binding partner IRF9, with STAT2 contributing a critical transactivation domain. Human RVB1 and RVB2, which are highly conserved AAA ؉ ATP binding proteins contained in chromatin-remodeling complexes such as Ino80, SNF2-related CBP activator protein (SRCAP), and Tip60/NuA4, interacted with the transactivation domain of STAT2 in the nuclei of IFN-stimulated cells. RNA interference (RNAi) experiments demonstrated that RVB proteins were required for robust activation of IFN-␣-stimulated genes (ISGs). The requirement for RVB proteins was specific to IFN-␣/ STAT2 signaling; transcription of tumor necrosis factor alpha (TNF-␣)-and IFN-␥-driven genes was not affected by RVB1 depletion. Using RNAi-based depletion, we assessed the involvement of catalytic subunits of the RVB-containing Tip60, BRD8, Ino80, SRCAP, and URI complexes. No component other than RVB1/2 was uniquely required for ISG induction, suggesting that RVB1/2 functions as part of an as yet unidentified complex. Chromatin immunoprecipitation assays indicated that RVB1/2 was required for recruitment of RNA polymerase II (Pol II) to ISG promoters but was dispensable for STAT2 recruitment to chromatin. We hypothesize that an RVB1/2 chromatin-remodeling complex is required for efficient Pol II recruitment and initiation at ISG promoters and is recruited through interaction with the STAT2 transactivation domain. Interferons (IFNs) are a family of pleiotropic cytokines primarily known for their ability to establish a potent antiviral state via modulation of the Janus kinase-signal transducer and activator of transcription (JAK-STAT) signal transduction pathway. Alternatively, aberrant production of IFNs correlates with systemic autoimmunity, underscoring the importance of the stringent regulation of IFN responses. Type I IFNs activate the heterotrimeric transcription factor ISGF3, composed of tyrosine-phosphorylated STAT1 and STAT2 and an auxiliary DNA binding protein, IRF9. Following IFN stimulation, phosphorylated ISGF3 translocates into the nucleus, binds the interferon-stimulated response elements (ISREs) in the promoters of IFN-␣/␤-stimulated genes (ISGs), and rapidly and robustly induces the transcription of a large family of previously silent genes. ISGs subsequently influence multiple cellular pathways involved in innate immunity in order to protect against viral and bacterial replication, including modulation of cell stress responses, apoptosis, proliferation, translation, and innate and adaptive immune signaling (1). In the context of ISGF3, STAT2 contains an essential transactivation domain (TAD) that provides most of the transcriptional function to the transcription factor complex, whereas STAT1 and IRF9 confer DNA sequence specificity (2). Induction of IFN target genes provides a robust system for studying the molecular mechanisms underlying transcriptional regulation. However, little is...

Suppression of Type I Interferon Signaling by E1A via RuvBL1/Pontin

“…Following IFN treatment, we observed considerable enrichment of E1A and RuvBL1 at the ISG56 and IFI6 promoters. Recruitment of RuvBL1 to these promoters was consistent with previous reports (24). Recruitment was specific to the promoter region, as we did not observe recruitment of either E1A or RuvBL1 to the 3= end of the ISG56 gene under either untreated or IFN-treated conditions (Fig.…”

“…ns, not significant. cating RuvBL1 in ISG regulation (24). Since these authors used 293 cells expressing wt E1A, it is difficult to interpret their results in light of our current findings.…”

“…Unexpectedly, we observed that depletion of RuvBL1 had an enhancing effect on ISG expression after IFN treatment, contrary to a previously published result (24). Although this was not significant for IFI6, it was for ISG56, and based on the earlier study, we expected some reduction in ISG expression after RuvBL1 depletion.…”

“…Our investigation of the role of RuvBL1 in viral replication showed only minimal effects on various measures of viral fitness, yet we observed a modest, but consistent, reduction in the ability of the virus to grow following RuvBL1 depletion. Previous reports have implicated RuvBL1 in regulation of ISGs, specifically ISG56 and IFI6 (24). We therefore investigated whether depletion of RuvBL1 affects the ability of HAdV, and more specifically E1A, to suppress ISG56 and IFI6 following infection.…”

Phosphorylation of STAT2 on serine-734 negatively regulates the IFN-α-induced antiviral response

“…In type I IFN signaling, this task is performed by STAT1 and IRF9, which bind DNA directly while STAT2 contributes its transactivation function to the ISGF3 complex. Therefore, we can postulate that the first part of the STAT2-TAD (from residues ∼700-760) is required to bring the remaining 90 residues (761-851) into a proper conformation for binding to co-activators such as p300/CBP, GCN5 and, most recently, RVB1 and RVB2 proteins (Gnatovskiy et al, 2013).…”