Chronic infection of hepatitis B virus (HBV) is closely associated with the development of human hepatocellular carcinoma (HCC). HBV X protein (HBx) plays a key role in the progression of HCC. We recently found that amplified in breast cancer 1 (AIB1) protein is overexpressed in 68% of human HCC specimens and promotes HCC progression by enhancing cell proliferation and invasiveness. Given that both HBx and AIB1 play important oncogenic roles in HCC, we aimed to determine whether they could cooperatively promote human HCC development. Herein, we show that HBx-positive HCC tissues had a higher level of AIB1 protein, compared to HBx-negative HCC tissues. A positive correlation between HBx protein level and AIB1 protein level was established in HCC specimens. Without affecting its messenger RNA level, HBx induced a significant increase of the protein level of AIB1, which correlated with a significant extension of the half-life of AIB1 protein. Mechanistically, HBx could interact with AIB1 to prevent the interaction between envelope protein 3 ubiquitin ligase F-box and WD repeat domain containing 7 (Fbw7)α and AIB1, then inhibited the Fbw7α-mediated ubiquitination and degradation of AIB1. In addition, reporter assays and chromatin immunoprecipitation assays revealed that both HBx and AIB1 were recruited to matrix metalloproteinase-9 (MMP-9) promoter to enhance MMP-9 promoter activity cooperatively. Consistently, HBx and AIB1 cooperatively enhanced MMP-9 expression in HepG2 cells, which, in turn, increased cell-invasive ability. Conclusion: Our study demonstrates that HBx can stabilize AIB1 protein and cooperate with it to promote human HCC cell invasiveness, highlighting the essential role of the cross-talk between HBx and AIB1 in HBV-related HCC progression. © 2012 American Association for the Study of Liver Diseases.