Research on The Regulatory Mechanism of Manganate Pathway to Enhance Plant Carbon Dioxide Conversion

Introduction

Terrestrial plants can convert atmospheric CO₂ into diverse and abundant aromatic compounds, thus providing high-value chemicals and considerable promise for carbon storage. However, it is unclear how plants control the shikimate pathway that connects the photosynthetic carbon fixation with the biosynthesis of aromatic amino acids, the major precursors of plant aromatic natural products. Understanding the regulatory mechanisms of the manganate pathway can be used to can be used to enhance plant-based, sustainable conversion of atmospheric CO₂ to high-energy and high-value aromatic compounds.

Fig. 1 Multiple suppressors of tyra2 (sota) mutations rescued the tyra2 growth inhibition and enhanced tyrosine (Tyr) and phenylalanine (Phe) accumulation.

Methods

Arabidopsis thaliana plants were placed under suitable growth conditions and cultured in soil supplied with Hoagland solution or on the agarose-containing 0.5-strength Murashige and Skoog medium with 1% sucrose. The seeds of the tyra2-1 transfer DNA insertion mutant were used to conduct a forward genetic suppressor screening using methanesulfonate (EMS). To identify the causal mutations leading to the suppression of the tyra2 phenotypes and the accumulation of aromatic amino acids (AAAs) in the metabolic sota lines, whole-genome sequencing and genetic mapping of approximately 200 seedlings displaying tyra2-like and sota-like phenotypes were performed, and the identified DHS sota mutations were validated. To further investigate the effect of sota mutation on plant DHS enzymes and the mechanism of regulation of the shikimate pathway by sota mutation, transgenic plant construction, DHS enzyme assay, differential scanning fluorescence analysis, soluble metabolite analysis, CO₂ labeling experiments, quantification of starch, sugar, protein and lignin contents, gas exchange measurements, fluorescence quantitative PCR expression analysis and amino acid sequence comparison experiments were carried out, respectively.

Results

Genetic analysis showed that all eight metabolic sota mutations examined mapped to loci encoding DHSs, but not to other shikimate pathway enzymes, suggesting that DHS-catalyzed reactions are important for the regulation of plant shikimate pathway. However, unlike microbial DHSs that are directly inhibited by the pathway product, AAAs, plant DHSs are subjected to highly complex feedback regulation mediated not only by AAAs but also by many AAA-derived compounds.

Fig. 2 The sota mutations biochemically deregulate the effector-mediated DHS negative feedback inhibition.

Furthermore, the excessive accumulation of aromatic amino acids and the increased photosynthesis observed in the sota mutant plants would provide additional energy to support the elevated activity of the highly energy-intensive shikimate pathway and AAA biosynthesis. Thus, our discovery of a series of DHS point mutations could provide powerful plant genetic tools to enhance the conversion of CO₂ into aromatic compounds for sustainable production of high-value chemicals while concomitantly reducing atmospheric CO₂.

Fig. 3 Carbon fixation is accelerated to support high AAA production in the sota mutants.

Summary

This study identified suppressor of tyra2 (sota) mutations that deregulate the first step in the plant shikimate pathway by alleviating multiple effector-mediated feedback regulation in Arabidopsis thaliana. The sota mutant plants showed hyperaccumulation of aromatic amino acids accompanied by up to a 30% increase in net CO₂ assimilation. The identified mutations can be used to enhance plant-based, sustainable conversion of atmospheric CO₂ to high-energy and high-value aromatic compounds.

Reference:

Yokoyama, R.; et al. Point mutations that boost aromatic amino acid production and CO₂ assimilation in plants. Science Advances, 2022, 8(23): eabo3416.

* For research use only. Not intended for any clinical use.

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Research on The Regulatory Mechanism of Manganate Pathway to Enhance Plant Carbon Dioxide Conversion

Introduction

Methods

Results

Summary

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