A PHOSPHONATE MONOESTER HYDROLASE FROM BURKHOLDERIA-CARYOPHILLI PG2982 IS USEFUL AS A CONDITIONAL-LETHAL GENE IN PLANTS

A bacterial phosphonate monoester hydrolase was evaluated in plants as a conditional lethal gene useful for cell ablation and negative selec tion. Glyphosate is a potent herbicide whereas its phosphonate monoest er derivative, glyceryl glyphosate, is approximately 50-fold less acti ve. A phosphonate monoesterase gene (pehA) encoding an enzyme that hyd rolyzes phosphonate esters including glyceryl glyphosate to glyphosate and glycerol was cloned from the glyphosate metabolizing bacterium, B urkholderia caryophilli PG2982. Constitutive expression of the pehA ge ne in Escherichia coil and Arabidopsis thaliana RLD had no observable phenotypic effects on growth and development. However, cells and plant s expressing the pehA gene were killed when treated with glyceryl glyp hosate. The phytotoxicity resulted from the hydrolysis of glyceryl gly phosate to glyphosate and subsequent inhibition of aromatic amino acid biosynthesis. As an example of tissue-specific cell ablation, floral sterility without vegetative toxicity was demonstrated by expressing t he pehA gene using a tapetal-specific promoter and treating the mature plants with glyceryl glyphosate. A chromogenic phosphonate ester subs trate, 5-bromo-4-chloro-indolyl phenylphosphonate, was used to monitor in situ expression of the pehA gene. The general utility of the pehA gene as a heterologous conditional lethal gene in plants is discussed.