Tissue transglutaminase (TG2), an enzyme that catalyzes Ca 2+-dependent aggregation and polymerization of proteins, is overexpressed in ovarian cancer cells and tumors. We previously reported that TG2 facilitates tumor dissemination using an i.p. xenograft model. Here we show that TG2 modulates epithelial-to-mesenchymal transition (EMT), contributing to increased ovarian cancer cell invasiveness and tumor metastasis. By using stable knockdown and overexpression in epithelial ovarian cancer cells, we show that TG2 induces a mesenchymal phenotype, characterized by cadherin switch and invasive behavior in a Matrigel matrix. This is mediated at the transcriptional level by altering the expression levels and function of several transcriptional repressors, including Zeb1. One mechanism through which TG2 induces Zeb1 is by activating the nuclear factor-κB complex. The effects of TG2 on ovarian cancer cell phenotype and invasiveness translate into increased tumor formation and metastasis in vivo, as assessed by an orthotopic ovarian xenograft model. Highly expressed in ovarian tumors, TG2 promotes EMT and enhances ovarian tumor metastasis by activating oncogenic signaling. ©2009 American Association for Cancer Research.