The innate immune system helps determine the outcome of pathogen invasion of a
host because it mediates recognition of and initiates the host response against the
pathogen. The balance of the system is delicate. Too little signaling is ineffective in
clearing the pathogen and too much is pathological for the host. Therefore, regulation is
important for both parties.
dsRNA is a molecular pattern associated with viral infections which stimulates
the innate immune response. Host recognition of dsRNA include Toll like receptor 3
(TLR3) and the RNA helicases retinoic acid inducible gene (RIG-I) and melanoma
differentiation associated gene (mda-5). These receptors signal to activate many
transcription factors including Interferon (IFN) Regulatory Factor 3 (IRF3) which
mediates the expression of IFN stimulated genes (ISGs). Of the hundreds of ISGs, ISG56
is consistently one of, if not the most, highly induced, indicating an importance in
antiviral immunity. Indeed, p56, the protein product of ISG56, inhibits host translation
and in doing so prevents viral replication.
To determine the physiological significance of p56, we sought to conditionally
disrupt the mouse ISG56 gene. (Chapter 3) Unfortunately, problems were encountered in
generating an embryonic stem cell containing a correctly integrated targeting construct.
To discover novel and to distinguish previously characterized regulatory
mechanisms of IRF3, we established a cell survival assay that can identify both activators
and inhibitors of IRF3 via TLR3 signaling. (Chapter 4) Using this system, we found that
V proteins encoded by select RNA Paramyxoviridae viruses could inhibit IRF3
activation by acting as alternative substrates for the IRF3 kinases inhibitor of κ B kinase
epsilon (IKKe) and TANK binding kinase (TBK1). (Chapter 5) In addition, we identified
novel open reading frames from the DNA viruses cytomegalovirus and murine
γ-herpesvirus 68 that could inhibit IRF3 activation. (Chapter 6) Mechanistic studies of
these inhibitors revealed complexities in IRF3 activation, distinguished TBK1 and IKKe
as points of convergence and divergence for many pathways and indicated the disparate
needs of viruses to both activate and inhibit immune signaling. (Chapter 7) These
investigations have implications not only in antiviral therapy development but also in
understanding host virus coevolution.