Journal of Energy Bioscience 2025, Vol.16, No.4, 205-215 http://bioscipublisher.com/index.php/jeb 210 6 Emerging Tools and Strategies 6.1 Synthetic biology approaches Synthetic biology provides a technical and theoretical basis for the modular reconstruction of the C4 photosynthetic pathway. By delving into the mechanism of action and evolutionary process of key enzymes in C4 metabolism, researchers have proposed a simplified engineering model based on known C4 metabolic components. These models have promoted the expression and functional verification of C4-related genes in C3 crops such as wheat. Furthermore, synthetic biology also supports methods such as targeted mutagenesis and directed evolution to optimize the activity and position of C4 enzymes in cells, thereby accelerating the engineering process of C4 traits (Schuler et al., 2016; Prasanna et al., 2025). 6.2 Advanced genome editing Advanced genome editing technologies such as CRISPR/Cas provide powerful tools for precisely regulating the functions of C4-related genes. Through gene knock-in, knockout and site-directed mutagenesis, the key C4 enzymes can be efficiently expressed in wheat leaves and localized to appropriate subcellular locations. These techniques can also be used to regulate factors related to the anatomical structure of Kranz, laying the foundation for the spatial partitioning required for C4 photosynthesis (Schuler et al., 2016; Cui, 2021; Prasanna et al., 2025). 6.3 Cell-type specific promoters The operation of the C4 photosynthetic pathway depends on the rational distribution of enzymes between mesophyll cells and vascular bundle sheath cells. In C4 plants, these functional genes have acquired new tissue expression patterns. Therefore, identifying and utilizing specific promoters has become a key step in achieving cell type-specific expression. By comparing genomes and analyzing promoter sequences, researchers have identified a variety of ccis elements that affect C4 gene expression and subcellular localization. These elements provide important molecular tools for wheat C4 engineering (Chen et al., 2023; Raturi et al., 2024). 6.4 Comparative systems biology Systems biology combines genomic, transcriptomic, proteomic and metabolomic data to help reveal the regulatory networks and evolutionary patterns of the C3 and C4 photosynthetic pathways. Comparative analysis reveals that C4-related genes already exist in C3 crops, but their expression patterns and localization still need to be optimized. Systems biology can also be used to simulate the effects of C4 engineering on the overall metabolism and growth of wheat, thereby providing references for multi-gene synergistic regulation and yield trait optimization (Schuler et al., 2016; Cui, 2021; Chen et al., 2023; Raturi et al., 2024). 7 Potential Impacts of Successful C4 Engineering in Wheat 7.1 Yield gains under different climates The engineering of the C4 photosynthetic pathway into wheat is expected to significantly increase yield under adverse conditions such as high temperature and drought. C4 plants (such as corn) have higher photosynthetic efficiency and more biomass accumulation under conditions of high temperature and water shortage. One important reason for this is that their CO2 fixation mechanism is more efficient (Rangan et al., 2016; Wang et al., 2022; Rezaei et al., 2023). Studies have found that wheat expresses some C4 photosynthetically related genes during the grain development stage. This indicates that optimizing the photosynthetic contributions of C3 and C4 through genetic improvement may enable wheat to have higher yield and more stable performance under extreme climates (Rangan et al., 2016). Simulation and field trial results show that if the wheat genotype has higher radiation utilization efficiency, earlier flowering time and larger grain weight, the yield can increase by 6% to 69% in the case of more frequent extreme weather in the future (Yan et al., 2021). In addition, if the source reservoir characteristics of wheat are improved, the yield can theoretically increase by 16% to 52% under current and future climatic conditions, but sufficient nitrogen fertilizer input is required (Martre et al., 2024). 7.2 Adaptation to climate change The C4 photosynthetic mechanism makes crops more heat-tolerant and drought-tolerant, which can alleviate the adverse effects brought about by climate change. Under the current climate change conditions, the yields of C3
RkJQdWJsaXNoZXIy MjQ4ODYzNA==