Chloroplast Engineering for Herbicide Resistance via Glutamine Synthetase

Introduction

Glutamine synthase (GS) plays a key role in plant metabolism. GS is the first and only enzyme capable of combining inorganic ammonium ions with organic compounds in the enzymatic tandem of the nitrogen uptake pathway responsible for glutamine synthesis. The isoenzyme GS2 of GS mainly exists in the plastids of green tissues and is mainly involved in the photorespiration release of NH3. This enzyme helps utilize ammonia produced by nitrite reductase and recycle ammonia produced by photorespiration. There are currently three main compounds that inhibit glutamine synthase, namely methionine sulfoxide (MSO), glufosinate and tabtoxinine-β-lactam. Inhibition of glutamine synthase activity results in rapid accumulation of ammonia, cessation of photosynthesis, disruption of chloroplast structure, and cytoplasmization of the substrate. Therefore, glutamine synthetase becomes the main target of herbicides.

Fig. 1. Schematic illustrating the basics of ammonium assimilation in photosynthetic cells and a summary of the reported effects of the absence of GS2. (Marino D, et al., 2022)

Solutions

Glutamate inhibits plant glutamine synthase and nitrogen metabolism, and aspartate tolerance has been engineered in more than 36 plants. Our engineers have been working to engineer herbicide resistance by encoding the GS2 gene, resulting in the successful development of glutonate-resistant crops. Here, Lifeasible introduced the GS2 gene into the chloroplast genome to confer highly efficient tolerance to glutonate without any deleterious effect on flowering or seed setting.

Glufosinate-ammonium (phosphinothricin, PPT), an analog of glutamate, is a non-selective broad-spectrum herbicide against GS. We offer a variety of crop-based glutamine synthase mutants to enhance glufosinate tolerance. Our available solutions for herbicide resistant via GS include:

• Glutamine synthase activity.
• Effects of glufosinate-ammonium on glutamate and glutamine levels.
• GS1a, GS1b and GS2 expression.
• Absorption and Transport of ¹⁴C-Glufosinate-ammonium.
• Metabolism of glufosinate.

Features of Our Strategy

• GS has a rapid response and very efficient, tight regulation at the substrate level, which can be achieved by allosteric.
• GS is involved in the primary assimilation of ammonia produced by nitrate reduction or nitrogen fixation in plant roots and the re-assimilation of ammonia released by photorespiration.
• PPT makes plants resistant to engineered herbicides, and ammonia accumulates rapidly, inhibiting photosynthesis and causing plant cell death.
• Gene amplification of GS enhances the expression of GS.
• Altering glutamate and glutamine concentrations in glufosinate-affected tissues.

GS2 is at the intersection of different processes such as photorespiration, nitrogen assimilation and stress tolerance. Breeding to alter the expression level of GS2 to confer glufosinate tolerance is a promising strategy. Lifeasible has extensive knowledge and experience in the engineering of herbicide resistance via herbicide-insensitive enzymes. Our mission is to provide customers with comprehensive, reliable, professional solutions to accelerate your research. If you are interested in our solutions, please contact us at any time.

References

1. Marino D, Cañas R A, Betti M. (2022) Is plastidic glutamine synthetase essential for C3 plants? A tale of photorespiratory mutants, ammonium tolerance and conifers[J]. New Phytologist. 234(5): 1559-1565.
2. Brunharo C A C G, Takano H K, Mallory-Smith C A, et al. (2019) Role of glutamine synthetase isogenes and herbicide metabolism in the mechanism of resistance to glufosinate in Lolium perenne L. spp. multiflorum biotypes from Oregon[J]. Journal of agricultural and food chemistry. 67(31): 8431-8440.