An Alternative Splicing Isoform of MITA Antagonizes MITA-Mediated Induction of Type I IFNs

Abstract

Mediator of IFN regulatory transcription factor 3 activation (MITA) is an important adaptor protein to mediate the induction of type I IFNs. In this study, we identified an alternatively spliced isoform of MITA lacking exon 7, termed MITA-related protein (MRP). MRP shares the N-terminal portion aa 1–253 with MITA but possesses a unique 30-aa sequence at the carboxyl terminal part, therefore lacking the conserved domains including TANK-binding kinase 1 (TBK1) and cyclic diguanylate binding domain. MRP is expressed in multiple tissues and distinct cell lines. Overexpression of MRP inhibited MITA-mediated activation of IFN-β promoter by sendai virus infection and cyclic diguanylate treatment but enhanced that in HSV-1 infection. Interestingly, MRP expression was reduced after Sendai virus infection but was upregulated after HSV-1 infection. Overexpression of MRP inhibited MITA-mediated induction of IFN-β via TBK1-IFN regulatory transcription factor 3 by disrupting the MITA-TBK1 interaction. However, NF-κB pathway was still activated by MRP, as MRP retained the ability to interact with inducible inhibitor of NF-κB (iκB) kinase. Thus, MRP acts as a dominant negative regulator of MITA-mediated induction of IFN production.

MITA/STING and Its Alternative Splicing Isoform MRP Restrict Hepatitis B Virus Replication

“…Although MRP was identified as a dominant negative mutant of MITA/STING, it retains the ability to activate NF-κB in 293T cells [29]. Therefore, it is worthwhile to investigate and compare the function of MITA/STING and MRP during HBV replication.…”

“…The plasmid was constructed by cloning the Eco RI head-to-tail dimer of HBV genome (genotype D), subtype ayw (GenBank accession number, V01460) into plasmid pMa5-8 [33, 34], using the authentic HBV promoter for HBV transcription. The human MITA/STING overexpression plasmid pFlag-MITA (pMITA), human MRP overexpression plasmid pHA-MRP (pMRP) and control vector plasmid pcDNA 3.1(+) were described previously [29]. The reporter plasmids pIFN-β-luc, pIRF3-luc, pNF-κB-luc, pISRE-luc were purchased from Clontech.…”

“…The siMRP was designed and had been proved to be specific to MRP [29]. The siRNAs were transfected using Lipofectamine RNAiMax (Invitrogen) at a final concentration of 80 nM referring to the manufacturer’s instructions.…”

“…In addition, MITA/STING also plays an important role in IFNs production during Dengue virus (DENV) infection [28]. An alternatively spliced variance of MITA/STING, designated as MITA-related protein (MRP), was found to act as a dominant negative mutant of MITA/STING, blocking MITA/STING-mediated IFN induction by disrupting the interaction between MITA/STING and TBK1; interestingly, MRP is still able to activate NF-κB [29]. Because dsDNA and ssDNA are produced during HBV replication, we propose that MITA and MRP are involved in the recognition of HBV replicative intermediates and the regulation of HBV clearance.…”

PLA1A Participates in the Antiviral Innate Immune Response by Facilitating the Recruitment of TANK-Binding Kinase 1 to Mitochondria

“…Mammalian expression plasmids for PLA1A, RIG-I, MAVS, STING, TBK1, IKKε, and IRF3 were constructed by standard molecular biology techniques as described previously [20,21].…”