Abstract

In addition to their role in detoxification, Glutathione S‐transferase (GST) enzymes influence signal transduction, development, and differentiation in many eukaryotic cells. Transcriptome analysis of the cellular slime mold, Dictyostelium discoideum (strain AX4) expresses five GST (α1‐α5 class) transcripts whose function(s) remain unknown. This study characterized the activity and expression of GST isozymes in D. discoideum at various stages of growth and development. A GSH‐agarose pull down assay of the lysates from vegetative amoebae identified two protein species of ~24kDa; mass spectrophotometric analysis identified GSTα3> GSTα1>GSTα2 as predominant species. Measurements of GST activity in amoeba using the substrate 1‐chloro‐2‐4‐dinitrobenzene (CDNB) revealed functional enzyme activity, which was significantly reduced by as much as 50% in the presence of 500 nM ethacrynic acid (EA, a GSTs inhibitor). Ethacrynic acid also delayed the formation of aggregates and multicellular development. Levels of GSTs activity differed significantly between vegetative amoebae and multicellular stages of development (aggregate, slug, culmination, fruiting body), indicating the importance of GSTs to influence cell signaling and gene expression during morphogenesis and differentiation. Current studies aim to determine: a) the levels of each isozyme and their substrate specificities; b) the impact of GST null mutants on growth and development of the organism. These studies will contribute to elucidating the expanding the role and function of GST enzymes in eukaryotic signal transduction that regulates cell growth, chemotaxis, and differentiation.

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