STRUCTURE AND REGULATION OF LIVER ACUTE PHASE GENES

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Acute phase proteins are produced by hepatocytes and are important in the defense against tissue damage and infections. Alpha2- macroglobulin (alpha2M), a broad range proteinase inhibitor, is increased several 100 fold-in acute and chronic inflammations; alpha-inhibitor III (alpha1I3), a close structural relative is strongly decreased. Transcription of both genes is controlled by glucocorticoids and interleukin 6 (IL6) in opposite directions. We have isolated and characterized DNA clones for both genes and have located a glucocorticoid responsive element in the alpha1I3 gene 5' flanking region. We have identified suitable rat hepatoma cell lines to study the regulation of both genes in culture and have also isolated rat IL6 DNA clones. These will be used to produce recombinant IL6 in order to map the IL6 responsive control elements of both genes. ln this continuation we will characterize the cis- and trans-acting control elements of both genes responsible for their regulation by these two hormones. The cis-elements will be mapped by transfection studies in hepatoma cells or transgenic mice. The elements responsible for hepatic transcription of both genes will be studied by cell free transcription in liver nuclear extracts. The unique availability of rat liver will be exploited to purify microgram amounts of relevant trans-factors by specific ORA affinity chromatography. Their function as transcription factors will be established by mutagenesis and complementation. Partial amino acid sequences of the factors will be determined and corresponding DNA clones will be isolated. Studies of the tissue specific and developmental regulation oF the factor genes will be initiated. Recombinant factors will be expressed in suitable systems and an initial characterization of their biochemical functions will be attempted. The purpose of this research is to characterize the elements mediating the transient control of acute phase gene transcription and to compare them with the elements responsible for the stable. liver specific expression of these and other genes. We wish to learn whether acute phase control is achieved by different combinations of the same elements that cause hepatic transcription or whether it involves qualitatively different elements. This is attempted by focusing on a system that is biochemically feasible, biologically relevant and already well characterized. Improved knowledge of gene regulation by inflammatory mediators may facilitate the future design of control substances of inflammation.