—Interleukin-6 (IL-6) is a multifunctional cytokine expressed by angiotensin II (Ang II)-stimulated vascular smooth muscle cells (VSMCs) that functions as an autocrine growth factor. In this study, we analyze the mechanism for Ang II-inducible IL-6 expression in quiescent rat VSMCs. Stimulation with the Ang II agonist Sar¹ Ang II (100 nmol/L) induced transcriptional expression of IL-6 mRNA transcripts of 1.8 and 2.4 kb. In transient transfection assays of IL-6 promoter/luciferase reporter plasmids, Sar¹ Ang II treatment induced IL-6 transcription in a manner completely dependent on the nuclear factor-κB (NF-κB) motif. Sar¹ Ang II induced cytoplasmic-to-nuclear translocation of the NF-κB subunits Rel A and NF-κB1 with parallel changes in DNA-binding activity in a biphasic manner, which produced an early peak at 15 minutes followed by a nadir 1 to 6 hours later and a later peak at 24 hours. The early phase of NF-κB translocation was dependent on weak simultaneous proteolysis of the IκBα and β inhibitors, whereas later translocation was associated with enhanced processing of the p105 precursor into the mature 50-kDa NF-κB1 form. Pretreatment with a potent inhibitor of IκBα proteolysis, TPCK, completely blocked Sar¹ Ang IIAng II-induced NF-κB activation and induction of endogenous IL-6 gene expression, which indicated the essential role of NF-κB in mediating IL-6 expression. We conclude that Ang II is a pleiotropic regulator of the NF-κB transcription factor family and may be responsible for activating the expression of cytokine gene networks in VSMCs.