Nuclear factor-kappaB (NF-kappaB) is a critical regulator of innate and adaptive immune function as well as cell proliferation and survival. The present study demonstrated for the first time that a virus belonging to the Arteriviridae family activates NF-kappaB in MARC-145 cells and alveolar macrophages. In porcine reproductive and respiratory syndrome virus (PRRSV)-infected cells, NF-kappaB activation was characterized by translocation of NF-kappaB from the cytoplasm to the nucleus, increased DNA binding activity, and NF-kappaB-regulated gene expression. NF-kappaB activation was increased as PRRSV infection progressed and in a viral dose-dependent manner. UV-inactivation of PRRSV significantly reduced the level of NF-kappaB activation. Degradation of IkappaB protein was detected late in PRRSV infection, and overexpression of the dominant negative form of IkappaBalpha (IkappaBalphaDN) significantly suppressed NF-kappaB activation induced by PRRSV. However, IkappaBalphaDN did not affect viral replication and viral cytopathic effect. PRRSV infection induced oxidative stress in cells by generating reactive oxygen species (ROS), and antioxidants inhibited NF-kappaB DNA binding activity in PRRSV-infected cells, suggesting ROS as a mechanism by which NF-kappaB was activated by PRRSV infection. Moreover, NF-kappaB-dependent expression of matrix metalloproteinase (MMP)-2 and MMP-9 was observed in PRRSV-infected cells, an observation which implies that NF-kappaB activation is a biologically significant aspect of PRRSV pathogenesis. The results presented here provide a basis for understanding molecular pathways of pathology and immune evasion associated with disease caused by PRRSV.