REDOX MODULATION OF THE EXPRESSION OF BACTERIAL GENES ENCODING CYSTEINE-RICH PROTEINS IN PLANT-PROTOPLASTS

Activity of neomycin phosphotransferase II (NPTII; gene, neo; five cys teines) in tobacco protoplasts transfected with fusions of the octopin e TR2' or cauliflower mosaic virus 35S promoter and the nea gene, with or without a signal peptide, increased up to 8-fold in response to ex ternally added dithiothreitol at concentrations that did not affect pr otoplast viability (up to 2.5 mM). Activity of phosphinothricin acetyl transferase (PAT; gene, bar; one cysteine) expressed under control of the TR1' or 35S promoter was not similarly affected, thus excluding a redox modulation of transcription as the mechanism of NPTII activation by dithiothreitol. Western blot analyses showed an increase in the am ount bf protein in response to dithiothreitol, whereas neither the ste ady-state level of NPTII mRNA nor the specific activity of the purifie d enzyme was affected. The same type of modulation was observed for tr ansiently expressed beta-glucuronidase (nine cysteines) produced from a fusion with the 35S promoter, with or without a signal peptide. Limi tation of cotranslational and/or early posttranslational steps by exce ssively oxidizing sulfhydryl/disulfide redox potentials is postulated to explain the low net accumulation of cysteine-rich proteins of bacte rial origin (i.e.; NPTII and beta-glucuronidase) when expressed in pla nt protoplasts, and the marked increase in such proteins in response t o externally added dithiothreitol.