Scientists Engineer Elite Herbicide-Tolerance Alleles by Coupling Protein Design with Gene-Expression Tuning

Weeds are among the greatest constraints on crop yield, and prolonged reliance on herbicides that target only a limited set of enzymes has driven the worldwide emergence of resistant weeds. Developing crops that tolerate new herbicides is therefore an important route to more effective weed control. Conventional genome editing, however, typically modifies either a gene’s protein-coding sequence or its regulatory region alone, making it difficult to improve a target trait without compromising the gene’s normal function.

Now, a research team led by Prof. GAO Caixia and Prof. CHEN Yuhang from the Institute of Genetics and Developmental Biology (IGDB) of the Chinese Academy of Sciences (CAS) has developed a “dual-layer” allele engineering strategy that overcomes this limitation. By coordinating structure-guided protein design with precise cis-regulatory tuning, the researchers generated rice and wheat with improved herbicide tolerance. The study was published in Nature Plants on October 12 (DOI: 10.1038/s41477-026-02423-4).

The team used rice 4-hydroxyphenylpyruvate dioxygenase (HPPD), an essential enzyme in tyrosine metabolism and a target of β-triketone herbicides, as a model. They determined the crystal structure of HPPD bound to the herbicide mesotrione and guided by the architecture of the binding pocket, designed and screened amino-acid substitutions that reduced herbicide sensitivity. Several of these coding-sequence variants raised resistance but also impaired the enzyme’s normal function and plant growth, revealing a trade-off between resistance and enzyme activity.

To resolve this trade-off, the researchers identified a 22 bp expression-modulating element (EME) and used prime editing to integrate it precisely into the regulatory region of the endogenous HPPD gene. Combining the EME with the coding-sequence variants enhanced herbicide tolerance synergistically. Some rice lines withstood mesotrione at four times the recommended field application rate, as well as other HPPD-inhibiting herbicides. Field trials further showed that the balance between herbicide tolerance and agronomic performance depended on the allelic configuration. Some combinations reduced seed setting and yield, whereas retaining one wild-type allele preserved high tolerance while balancing it with agronomic traits.

The team then extended the strategy to allohexaploid wheat, editing the coding and regulatory regions of wheat HPPD simultaneously to obtain lines carrying both a coding variant and the EME. These lines showed elevated HPPD expression and stronger tolerance to mesotrione than wild-type plants, demonstrating that the approach is effective even in crops with complex genomes.

By optimizing protein function and gene dosage together, the dual-layer strategy can reach beneficial alleles that natural variation, conventional mutagenesis, or single-layer editing rarely deliver. Because the EME is compact, introduces no foreign gene cassette, and relies on sequence motifs already common in crop genomes, it combines a low regulatory footprint with predictable control of expression.

More broadly, the work provides a general framework for engineering dosage-sensitive agronomic traits, in which a gain in one function must be balanced against an essential biological role. It offers an effective strategy and technical route for the precise improvement of complex crop traits and for sustainable weed management.

Contact:

Prof. GAO Caixia

Institute of Genetics and Developmental Biology, Chinese Academy of Sciences

Email: cxgao@genetics.ac.cn