Inhibition of the dsRNA Response by Picornaviruses

The dsRNA response is a critical defense pathway used by cells to guard against viral infection. Critical
for the function of this pathway is the production of IFN-p, which requires the activation of the latent
transcription factor IRF-3. IRF-3 is normally found in the cytoplasm, but upon exposure to dsRNA or viral
infection IRF-3 is phosphorylated, dimerizes and translocates into the nucleus where it activates transcription
of IFN-p. Recently, we found that infection of HeLa cells with rhinovirus type 14 (RV14) prevents dimerization
of IRF-3 and production of IFN-b, and thus attenuates this response. The overall goal of this proposal is to
determine the underlying mechanisms responsible for inhibition of the dsRNA response by rhinovirus and
determine if other picornaviruses utilize a similar strategy to evade the host innate immune response. The
first aim of this proposal will examine the mechanism responsible for the lack of IFN-p mRNA and IRF-3
dimerization in rhinovirus-infected cells. This will be accomplished by determining the rate of transcription
from the IFN-p promoter using nuclear run-on experiments and by determining if phosphorylation of IRF-3 is
disrupted in infected cells. The second aim of this proposal will identify the viral factors responsible for
inhibition of the dsRNA response by examining induction of the dsRNA response in cell lines that express
individual viral proteins. Mutants of identified proteins will be generated and analyzed to determine the
functional domains responsible for inhibition of the dsRNA response. Ultimately, these mutations will be
introduced into an infectious rhinovirus cDNA to determine the role that inhibition of the dsRNA response has
in viral replication. The third aim will determine if inhibition of the dsRNA response extends to other cellular
promoters and other picornaviruses. Full genome Affymetrix microarrays will be used to determine the global
host transcriptional response to dsRNA in mock or RV14-infected cells. Subsequently, we will determine if
activation and attenuation of the dsRNA response is unique to rhinoviruses or extends to other
picornaviruses. This will be accomplished by examining IRF-3 and NF-icB activation and the global
transcriptional response in cells infected with other picornaviruses.
These studies will enhance our understanding of how rhinovirus evades host immune defense
mechanisms and may lead to the development of better therapeutics and strategies to combat these
prevalent human pathogens. In addition, these experiments will also provide us with general insights into the
basic cellular processes of signal transduction and transcriptional regulation. With this understanding it may
be possible to counteract the effect of infectious agents and reduce disease and mortality in humans,
animals and plants.