Self Protection from Anti-Viral Responses – Ro52 Promotes Degradation of the Transcription Factor IRF7 Downstream of the Viral Toll-Like Receptors

Rowan Higgs,Joan Ní Gabhann,Alexander Espinosa,Caroline A Jefferies,Claire Wynne,Marie Wahren-Herlenius,Elisa Lazzari,Derya Unutmaz

doi:10.1371/journal.pone.0011776

Rowan Higgs, Joan Ní Gabhann + Show 6 more

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https://doi.org/10.1371/journal.pone.0011776

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Abstract

Ro52 is a member of the TRIM family of single-protein E3 ligases and is also a target for autoantibody production in systemic lupus erythematosus and Sjögren's syndrome. We previously demonstrated a novel function of Ro52 in the ubiquitination and proteasomal degradation of IRF3 following TLR3/4 stimulation. We now present evidence that Ro52 has a similar role in regulating the stability and activity of IRF7. Endogenous immunoprecipitation of Ro52-bound proteins revealed that IRF7 associates with Ro52, an effect which increases following TLR7 and TLR9 stimulation, suggesting that Ro52 interacts with IRF7 post-pathogen recognition. Furthermore, we show that Ro52 ubiquitinates IRF7 in a dose-dependent manner, resulting in a decrease in total IRF7 expression and a subsequent decrease in IFN-α production. IRF7 stability was increased in bone marrow-derived macrophages from Ro52-deficient mice stimulated with imiquimod or CpG-B, consistent with a role for Ro52 in the negative regulation of IRF7 signalling. Taken together, these results suggest that Ro52-mediated ubiquitination promotes the degradation of IRF7 following TLR7 and TLR9 stimulation. As Ro52 is known to be IFN-inducible, this system constitutes a negative-feedback loop that acts to protect the host from the prolonged activation of the immune response.

Highlights

A critical step in the innate immune response to viral infection is the production of type I IFN (IFN-a and -b) [1]
Ro52 associates endogenously with IRF7 We have previously shown that recombinant Ro52 interacts with both overexpressed IRF3 and IRF7 in a pulldown experiment [13]
In order to confirm the association of Ro52 with endogenous IRF7 and to examine the status of this relationship following TLR stimulation, HeLa cells were stimulated with either polyI:C, lipopolysaccharide (LPS), imiquimod or CpG-B for 4 and 18 hours

Summary

Introduction

A critical step in the innate immune response to viral infection is the production of type I IFN (IFN-a and -b) [1]. The ubiquitin E3 ligase TNF receptor-associated factor 6 (TRAF6) is known to ubiquitinate IRF7 in a K63-dependent manner following TLR stimulation, resulting in IRF7 activation and the initiation of IFN-a transcription [14]. This effect has been shown to depend on the formation of a complex between IRF7, TRAF6 and MyD88 and is critical for type I IFN production downstream of TLR7, TLR8 and TLR9 stimulation [14,15]. The E3 ligase activity of TRAF6 is known to be required for latent membrane protein 1 (LMP1)-stimulated IRF7 ubiquitination and may be a prerequisite for IRF7 phosphorylation by IKKa, IKKe, TANK-binding kinase 1, or IL-1 receptor-associated kinase 1

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