The U937 human promyelocytic leukemia cell line has provided a valuable model for studying the mechanism of action of established and prospective treatment modalities in the laboratory. The U937 cell model has been used extensively for examining the mechanisms mediating the cytotoxic effects of chemotherapeutic agents as well as other forms of genotoxic stress, including UV radiation. Recent findings suggest that DNA damaging chemotherapeutics and UV radiation elicit their cytotoxic effects by triggering a sequence of events referred to as programmed cell death or apoptosis. Furthermore, the recently characterized protein cascade, termed the stress-activated protein kinase (SAPK) signal transduction pathway, has been implicated in mediating this response. To address this issue, a U937 cell model has been established in which the conditional expression of the dual-specificity protein phosphatase, MKP-1, functionally inhibits activation of the stress- activated signaling pathway. Preliminary studies indicate that conditional expression of MKP-1 provides cytoprotection against UV radiation-induced apoptosis. Thus, we have defined a novel anti- apoptotic role for MKP-1 in mediating cell survival in response to genotoxic stress stimuli. These findings provide the basis for a molecular model inwhich physiological resistance to genotoxic stress- induced apoptosis may be mediated by the induction of MKP-1, which attenuates signaling through the stress-activated protein kinase pathway. It is the goal of this proposal to determine the biochemical and genetic events regulating MKP-1-mediated resistance to genotoxic stress-induced apoptosis in the U937 cell model. It is our hypothesis that MKP-1 provides cytoprotection against genotoxic stress-induced apoptosis in U937 cells by attenuating the activation of SAPK or a SAPK- like protein kinase. Further, we hypothesize that this might provide a molecular basis for cellular resistance to chemotherapy.
