The importance of NF-kappaB activation and deficient anti-viral interferon induction in the pathogenesis of rhinovirus-induced asthma exacerbations is poorly understood. We provide the first in vivo evidence in man and mouse that rhinovirus infection enhanced bronchial epithelial cell NF-kappaB p65 nuclear expression, NF-kappaB p65 DNA binding in lung tissue and NF-kappaB-regulated airway inflammation. In vitro inhibition of NF-kappaB reduced rhinovirus-induced pro-inflammatory cytokines but did not affect type I/III interferon induction. Rhinovirus-infected p65-deficient mice exhibited reduced neutrophilic inflammation, yet interferon induction, antiviral responses and virus loads were unaffected, indicating that NF-kappaB p65 is required for pro-inflammatory responses, but redundant in interferon induction by rhinoviruses in vivo. Conversely, IFNAR1(-/-) mice exhibited enhanced neutrophilic inflammation with impaired antiviral immunity and increased rhinovirus replication, demonstrating that interferon signalling was critical to antiviral immunity. We thus provide new mechanistic insights into rhinovirus infection and demonstrate the therapeutic potential of targeting NF-kappaB p65 (to suppress inflammation but preserve anti-viral immunity) and type I IFN signalling (to enhance deficient anti-viral immunity) to treat rhinovirus-induced exacerbations of airway diseases.