Adenosine monophosphate-activated protein kinase (AMPK) is a metabolic regulator that promotes energy conservation and restoration when cells are exposed to nutrient stress. Given the high metabolic requirement of cancer cells, AMPK activation has been suggested as a potential preventative and therapeutic target. However, previous findings have shown that AMPK activity is diminished in some cancers. Expression of the 2 catalytic isoforms, AMPKa1 and AMPKa2, was evaluated in primary breast cancer and matched nontumor-adjacent tissue samples using immunohistochemistry. AMPK-dependent growth signaling events were examined in primary human mammary epithelial cells (HMECs) using RNAi to understand the importance of AMPKa2 in normal growth regulation. To test whether AMPKa2 would reinstate growth control and apoptotic mechanisms in breast cancer cells, metabolic stress assays and tumor xenografts were performed in MCF-7 cells, expressing low levels of AMPKa2, with stable transfection of either green fluorescent protein (GFP) or AMPKa2 expression constructs. AMPKa2 was found to be significantly suppressed in breast cancer tissue samples, whereas AMPKa1 was not. In normal HMECs, low glucose stress resulted in AMPK-driven growth inhibition. Interestingly, this response was ablated when AMPKa2 was silenced. Metabolic stress assays in MCF-7 cells indicated that AMPKa2 expression reduced both mTOR signaling and cyclin D1 expression, contributing to G1-phase cell cycle arrest. Cells expressing AMPKa2 underwent apoptosis more readily than GFP control cells. Xenograft studies demonstrated that MCF-7 tumors expressing AMPKa2 display reduced proliferation and increased apoptotic events. Furthermore, AMPKa2 xenografts exhibited diminished cyclin D1 levels along with an increased amount of nuclear p53, thereby implicating the AMPKa2-p53 signaling axis as a mediator of cell apoptosis. Together, these results highlight the significance of reduced AMPK activity contributing to human carcinogenesis and, specifically, the role of AMPKa2 with respect to its control of normal mammary epithelial cell growth and its reduced expression in breast cancer.