Small molecules of plant origin offer presumptively safe opportunities to prevent carcinogenesis, mutagenesis and other forms of toxicity in humans. However, the mechanisms of action of such plant-based agents remain largely unknown. In recent years the stress responsive transcription factor Nrf2 has been validated as a target for disease chemoprevention. Withania somnifera (WS) is a herb used in Ayurveda (an ancient form of medicine in South Asia). In the recent past, withanolides isolated from WS, such as Withaferin A (WA) have been demonstrated to be preventive and therapeutic against multiple diseases in experimental models. The goals of this study are to evaluate withanolides such as WA as well as Withania somnifera root extract as inducers of Nrf2 signaling, to probe the underlying signaling mechanism of WA and to determine whether prevention of acetaminophen (APAP)-induced hepatic toxicity in mice by WA occurs in an Nrf2-dependent manner. We observed that WA profoundly protects wild-type mice but not Nrf2-disrupted mice against APAP hepatotoxicity. WA is a potent inducer of Nrf2-dependent cytoprotective enzyme expression both in vivo and in vitro. Unexpectedly, WA induces Nrf2 signaling at least in part, in a Keap1-independent, Pten/Pi3k/Akt-dependent manner in comparison to prototypical Nrf2 inducers, sulforaphane and CDDO-Im. The identification of WA as an Nrf2 inducer that can signal through a non-canonical, Keap1-independent pathway provides an opportunity to evaluate the role of other regulatory partners of Nrf2 in the dietary and pharmacological induction of Nrf2-mediated cytoprotection.