Methionine adenosyltransferase (MAT) is an essential enzyme required for S‐adenosylmethionine biosynthesis. Hepatic MAT activity falls during chronic liver injury, and mice lacking Mat1a develop spontaneous hepatocellular carcinoma by 18 months. We have previously demonstrated that CD133⁺CD45⁻ oval cells isolated from 16‐month‐old Mat1a^−/− mice represent a liver cancer stem cell population. The transforming growth factor β (TGF‐β) pathway constitutes a central signaling network in proliferation, apoptosis, and tumorigenesis. In this study, we tested the response of tumorigenic liver stem cells to TGF‐β. CD133⁺CD45⁻ oval cells were isolated from premalignant 16‐month‐old Mat1a^−/− mice by flow cytometry and expanded as five clone lines derived from a single cell. All clone lines demonstrated expression of both hepatocyte and cholangiocyte markers and maintained a small population (0.5% to 2%) of CD133⁺ cells in vitro, and three of five clone lines produced tumors. Although TGF‐β1 inhibited cell growth equally in CD133⁻ and CD133⁺ cells from each clone line, the CD133⁺ population demonstrated significant resistance to TGF‐β–induced apoptosis compared with CD133⁻ cells. Furthermore, CD133⁺ cells demonstrated a substantial increase in mitogen‐activated protein kinase (MAPK) pathway activation, as demonstrated by phosphorylated extracellular signal‐regulated kinase levels before and after TGF‐β stimulation. MAPK inhibition using mitogen‐activated protein kinase kinase 1 (MEK1) inhibitor PD98059 led to a significant increase in TGF‐β–induced apoptosis in CD133⁺ cells. Conversely, a constitutively active form of MEK1 blocked the apoptotic effects of TGF‐β in CD133⁻ cells. Conclusion: CD133⁺ liver cancer stem cells exhibit relative resistance to TGF‐β–induced apoptosis. One mechanism of resistance to TGF‐β–induced apoptosis in CD133⁺ cancer stem cells is an activated mitogen‐activated protein kinase/extracellular signal‐regulated kinase pathway. (HEPATOLOGY 2009.)