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Adenovirus vectors (Adv) elicit innate immune responses via several pattern-recognition receptors. Although it has been suggested that various Adv-induced mechanisms play important roles in the induction of innate immunity in vitro , the impacts of these mechanisms in vivo remain unclear. Viral nucleic acids elicit innate immune responses through the recognition of cytosolic nucleic acid sensors and transduce intracellular signals to TANK-binding kinase (TBK) 1. In this study, to determine the impacts of viral nucleic acids on innate immune responses in vivo , we administered transgene-expressing Adv to Tbk1 -deficient mice. The systemic Adv administration failed to induce type I interferons (type I IFNs) in the spleen, but not the liver, of Tbk1 -deficient mice, resulting in the increase of transgene-expressing cells in the spleen, but not the liver. Moreover, Adv failed to induce type I IFNs in the bone-marrow-derived dendritic cells, but not the mouse embryonic fibroblasts, from Tbk1 -deficient mice in vitro . These results support the idea that Adv elicit innate immunity in immune cells and non-immune cells in a TBK1-dependent and TBK1-independent manner, respectively.