Aldo-keto Reductase Metabolizes Glyphosate and Confers Glyphosate Resistance in <i>Echinochloa colona</i>

Abstract

Glyphosate, the most commonly used herbicide in the world, controls a wide range of plant species, mainly because plants have little capacity to metabolize (detoxify) glyphosate. Massive glyphosate use has led to world-wide evolution of glyphosate-resistant (GR) weed species, including the economically damaging grass weed <i>Echinochloa colona</i> An Australian population of <i>E</i> <i>colona</i> has evolved resistance to glyphosate with unknown mechanisms that do not involve the glyphosate target enzyme 5-enolpyruvylshikimate-3-P synthase. GR and glyphosate-susceptible (S) lines were isolated from this population and used for resistance gene discovery. RNA sequencing analysis and phenotype/genotype validation experiments revealed that one aldo-keto reductase (AKR) contig had higher expression and higher resultant AKR activity in GR than S plants. Two full-length <i>AKR</i> (<i>EcAKR4</i>-<i>1</i> and <i>EcAKR4</i>-<i>2</i>) complementary DNA transcripts were cloned with identical sequences between the GR and S plants but were upregulated in the GR plants. Rice (<i>Oryza sativa</i>) calli and seedlings overexpressing <i>EcAKR4</i>-<i>1</i> and displaying increased AKR activity were resistant to glyphosate. EcAKR4-1 expressed in <i>Escherichia coli</i> can metabolize glyphosate to produce aminomethylphosphonic acid and glyoxylate. Consistent with these results, GR <i>E</i> <i>colona</i> plants exhibited enhanced capacity for detoxifying glyphosate into aminomethylphosphonic acid and glyoxylate. Structural modeling predicted that glyphosate binds to EcAKR4-1 for oxidation, and metabolomics analysis of <i>EcAKR4</i>-<i>1</i> transgenic rice seedlings revealed possible redox pathways involved in glyphosate metabolism. Our study provides direct experimental evidence of the evolution of a plant AKR that metabolizes glyphosate and thereby confers glyphosate resistance.