The glutathione S-transferases (GSTs) are a multi-class family of enzymes found in the cytosol of most cells. GSTs are enzymes of detoxication and protect cells from injury by a variety of metabolites and carcinogens. Most recently the GSTs in rodents have been found to function as inhibitors of a MAP kinase that is important in the cellular response to oxidative stress and homeostatic gene regulation. This proposal describes experiments that will determine the role of GSTs in signal transduction within human cells. The studies will be performed using a human gastric cancer cell line, Kato III, and an imortalized human stellate cell line, LX-2. These cell lines have been selected because they are proliferative and have the form of GST, termed P1-1, that is an inhibitor of the MAP kinase c-Jun-N-terminal kinase (JNK). Experiments will be performed to characterize the interaction between JNK and GSTP1-1. RNA interference technology will be used to inhibit expression of GSTP1-1 in both cell lines and the effect on JNK activity will be determined. The cell lines will also be subjected to an oxidative stress and the effects on JNK activity and proliferation will be determined using cells that do and do not express GSTP1-1. Lastly, the effect of the loss of GSTP1-1 expression on the susceptibility of the cells to apoptosis will be determined. From these studies we will define a role for GSTP1-1 in the regulation of gene expression, proliferation, and apoptosis via its effects on JNK activity in human cells. We have observed that the cytokine interleukin 6 and the acute phase response lead to a decrease in the expression of an important alpha class form of GST in rat liver. We have defined the region in the promoter of this gene that mediates the effect and have cloned a nuclear protein that is a negative regulator of GST and albumin expression during the acute phase response. We will further characterize this protein and determine how it functions as a negative regulator of gene expression in rat and human cells. The studies will clarify why expression of certain proteins fall during the acute phase response.
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