Overexpression of improvedEPSPSgene results in field level glyphosate tolerance and higher grain yield in rice

Glyphosate is a popular, systemic, broad-spectrum herbicide used in modern agriculture. Being a structural analog of phosphoenolpyruvate (PEP), it inhibits 5-enolpyruvylshikimate 3-phosphate synthase (EPSPS) which is responsible for the biosynthesis of aromatic amino acids and various aromatic secondary metabolites. Taking a lead from glyphosate-resistant weeds, two mutant variants of the riceEPSPSgene were developed by amino acid substitution (T173I + P177S;TIPS-OsEPSPSand G172A + T173I + P177S;GATIPS-OsEPSPS). These mutatedEPSPSgenes were overexpressed in rice under the control of either native EPSPS or constitutive promoters (maize ubiquitin [ZmUbi] promoter). The overexpression ofTIPS-OsEPSPSunder the control of theZmUbipromoter resulted in higher tolerance to glyphosate (up to threefold of the recommended dose) without affecting the fitness and related agronomic traits of plants in both controlled and field conditions. Furthermore, such rice lines produced 17%-19% more grains compared to the wild type (WT) in the absence of glyphosate application and the phenylalanine and tryptophan contents in the transgenic seeds were found to be significantly higher in comparison with WT seeds. Our results also revealed that the native promoter guided expression of modifiedEPSPSgenes did not significantly improve the glyphosate tolerance. The present study describing the introduction of a crop-specific TIPS mutation in class IaroAgene of rice and its overexpression have potential to substantially improve the yield and field level glyphosate tolerance in rice. This is the first report to observe that the EPSPS has role to play in improving grain yield of rice.