Reactive oxygen species (ROS) have been closely associated with cellular transformation, and promoting and sustaining tumorigenesis.  Elevated ROS levels have been shown to trigger opening of the mitochondrial permeability transition pore (MPTP), which can lead to mitochondrial dysfunction and ultimately, cell death.  In cancer cells the MPTP appears to be less susceptible to opening, which may account for their ability to proliferate in a more oxidative environment.  It is well established that signal transducer and activator of transcription 3 (STAT3), is constitutively activated in numerous human cancers. Recent studies indicate that STAT3 is also present in the mitochondria where it controls the activity of the electron transport chain, the production of ROS, Ras transformation of cells and growth of breast cancer cells. We now provide evidence that in response to oxidative insult, mitochondrial STAT3 interacts with Cyclophilin D (CypD), the key regulator and activator of the MPTP. Treatment of a variety of cells with hydrogen peroxide (H₂0₂) induces a rapid cycling of mitochondrial STAT3 where we have observed both its loss and re-accumulation in the mitochondria. We hypothesize that binding of CypD with STAT3 protects against oxidative stress induced cell death in cancer cells by preventing activation of the MPTP. We also provide evidence that mitochondrial STAT3 may play an important role in cancer cell resistance to chemotherapy. Consistent with this possibility, we observe a robust and selective increase in mitochondrial STAT3 levels following chemotherapeutic treatment of cancer cells. While prior reports have focused on the nuclear function of STAT3 in mediating tumorigenesis and therapeutic resistance, these results suggest a previously unappreciated and analogous role of STAT3 in the mitochondria.