Plasmacytoid dendritic cells (pDCs) are characterized by their ability to produce high levels of type 1 interferons in response to ligands that activate TLR7 and TLR9, but the signaling pathways required for IFN production are incompletely understood. Here we exploit the human pDC cell line Gen2.2 and improved pharmacological inhibitors of protein kinases to address this issue. We demonstrate that ligands that activate TLR7 and TLR9 require the TAK1-IKKbeta signaling pathway to induce the production of IFNbeta via a pathway that is independent of the degradation of IkappaBalpha. We also show that IKKbeta activity, as well as the subsequent IFNbeta-stimulated activation of the JAK-STAT1/2 signaling pathway, are essential for the production of IFNalpha by TLR9 ligands. We further show that TLR7 ligands CL097 and R848 fail to produce significant amounts of IFNalpha because the activation of IKKbeta is not sustained for a sufficient length of time. The TLR7/9-stimulated production of type 1 IFNs is inhibited by much lower concentrations of IKKbeta inhibitors than those needed to suppress the production of NFkappaB-dependent proinflammatory cytokines, such as IL-6, suggesting that drugs that inhibit IKKbeta may have a potential for the treatment of forms of lupus that are driven by self-RNA and self-DNA-induced activation of TLR7 and TLR9, respectively.