The glutathione S-transferases (GSTs) comprise a multi-class family of enzymes found in the cytosol of most cells. GSTs are enzymes of detoxification and protect cells from injury caused by a variety of endogenous and exogenous toxins and carcinogens. GSTs detoxify these reactive molecules by forming glutathione conjugates. The GSTs are dimeric proteins with independent catalytic sites, and it is unclear how the dimeric structure is necessary for function. The GST genes contain a number of regulatory elements that mediate increased expression in response to drugs. The physiologic factors that mediate expression of GSTs in the liver are undefined. Finally, different cells express different GST isozymes and the mechanisms that account for this cell specific expression are unknown. This proposal describes experiments that examine the affect of subunit interactions on catalysis. Techniques have been developed that allow the formation of heterodimers with mutations in only one of the two subunits. This approach is unique and allows for studies not previously possible. it was observed that the cytokine, interleukin 6, decreases the expression of a number of GST isozymes. The decrease in expression is mediated by elements in the promoter sequence of the GST gene and a unique nuclear protein has been identified that appears to mediate the decrease in transcription. This nuclear protein also is induced in the livers of animals during the acute phase response, and its appearance is associated with a decline in GST expression. In this proposal experiments are described that will further characterize the nuclear protein and define how it leads to a decrease in rates of transcription. Hepatic stellate cells (HSCs) upon activation become collagen-producing cells and account for the majority of collagen present in the cirrhotic liver. Normal HSCs express numerous GST isozymes, but upon activation most GSTs are lost and enzymatic activity against products of oxidant stress is reduced. Studies are proposed that will examine whether restoration of GST expression in activated HSC is associated with increased protection against oxidant injury and less collagen production in the GST expressing cells. It will also be determined whether the promoter region of the GST gene mediates the loss of expression of GST in activated HSCs. Finally, it has been observed that drugs such as phenobarbital induce GST expression in the liver both by increasing rates of transcription and mRNA stability. The latter appears to account for the greater expression of GSTs in pericentral vs periportal hepatocytes following treatment with phenobarbital. How phenobarbital selectively stabilizes mRNA in perivenous hepatocytes will be examined further.
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