A growing global population and the increasing prevalence of diet-related health issues such as “hidden hunger”, obesity, hypertension, and diabetes necessitate a fundamental rethinking of crop design and breeding. Synthetic metabolic engineering offers a method to modify and redesign metabolic pathways to increase the nutritional value of crops. We summarize recent advances in the biofortification of key nutrients including provitamin A, vitamin C, vitamin B9, iron, zinc, anthocyanins, flavonoids, and unsaturated fatty acids. We discuss the potential of multi-gene stacking, gene editing, enzyme engineering, and artificial intelligence in synthetic metabolic engineering. We propose future research directions and potential solutions centered on leveraging AI-driven systems biology, precision gene editing, enzyme engineering, agrobacterium-mediated genotype-independent transformation, and modular metabolic engineering strategies to develop next-generation nutritionally enhanced super crops and transform global food systems.

Hybrid sterility limits the use of strong interspecific heterosis and S19 is a major locus that confers hybrid sterility between Oryza sativa (Asian cultivated rice) and Oryza glaberrima (African cultivated rice). However, the S19 is not yet cloned and its underlying mechanism remains elusive. In this study, we identify two closely linked genes (S19A1 and S19A7) specific to African rice allele that encode a killer-protector module at the S19 locus. Two alternatively spliced transcripts expressed from the killer gene S19A1 (S19A1.1 and S19A1.2) encode mitochondria-targeted cytotoxic proteins that cause toxicity diversity for somatic and/or gametic cell death, respectively. However, S19A7 interacts with S19A1.1 and S19A1.2, blocking their cytotoxic effect. Because the Asian rice S19 allele lacks S19A1 and S19A7, male gametes that carry this allele are selectively aborted in Asian-African F1 hybrids. Knockout of S19A1 can overcome S19-mediated hybrid sterility. Haplotype analysis reveals that the functional S19 allele is absent in non-AA-genome Oryza species and likely emerged in the O. barthii-O. glaberrima lineage through a multi-step evolutionary process. Our findings provide insight into the genetic mechanisms responsible for hybrid sterility between Asian and African rice and suggest genetic and biotechnological strategies for the use of interspecific heterosis in rice.

Rice is one of the most important food crops and global warming, and extreme high temperatures severely threaten rice production. Rice is highly sensitive to heat stress during the reproductive stage, resulting in pollen sterility, a reduced seed-setting rate, and poor grain quality. Therefore, identifying key genes and elucidating the molecular mechanisms responding to heat stress can be leveraged to develop climate-resilient rice varieties, ultimately enhancing adaptation to the impact of future climate change on global rice production. Considerable progress has been made in elucidating the mechanisms of heat stress in rice. This review summarizes recent advances in our understanding of the molecular mechanisms underlying heat tolerance in rice, with a focus on the reproductive stage, including spikelet development, pollen fertility, pistil development, diurnal flower-opening time, grain filling, and grain quality. Additionally, we propose integrated strategies for mitigating high-temperature damage, addressing persistent challenges, and outlining future research directions for breeding heat-tolerant rice varieties.

Exo-Cas12i2 v1, a fusion of the 5' exonucleases T5E and PapE, facilitates editing of TA-rich regions and mediates deletions of large genomic fragments. Exo-Cas12i2 v1-driven MITE manipulation enables precise regulation of genes involved in gibberellin-mediated cell elongation and root ethylene responses to generate favorable agronomic traits.

Plant pectate lyases remodel the pectin matrix to drive cell expansion during plant growth and development. Here we show that OsPSE1, originally identified through the rice premature-senescence mutant ospse1, encodes an active pectate lyase that governs root pectin homeostasis and plant senescence. OsPSE1 is highly expressed in the root, where the wild-type OsPSE1 protein localizes primarily to the plasma membrane of root cells; however, the mutant ospse1 protein localizes almost exclusively to the nucleus. An in vitro depolymerase assay and quantitative tests showed that recombinant OsPSE1 protein has pectate lyase activity and can catalyze the degradation of polygalacturonic acid; the ospse1 protein shows reduced activity. Furthermore, ruthenium red staining, immunolabeling, and quantitative analysis revealed that the dysfunction of OsPSE1 causes abnormal accumulation of pectin and methyl-esterified homogalacturonan in the cell wall of rice root. Intriguingly, overexpression of OsPSE1 exhibits a significantly prolonged vegetative stage, compared to the wild type, with an extended reproductive stage and a longer overall growth period, which may be due to reduced pectin accumulation of roots and enhanced nutrient uptake in roots. Our findings indicate that the root pectin degradation regulated by the pectate lyase OsPSE1 is involved in rice plant senescence.

This commentary summarizes hybrid sterility models in plants, with an emphasis on a recent study that addresses the genetic basis of the RIS/RIA-RID-RIR system underlying S44-mediated hybrid sterility between Oryza longistaminata and indica rice, revealing a novel killer-protector-target model integrated with modifiers that regulate reproductive isolation.

Betalain, an economically valuable water-soluble natural plant pigment, is prized for its strong antioxidant activity, making it popular as a dietary supplement and a visual marker for plant transformation. However, market demand significantly outstrips current production capacity. This study reports the development of an efficient push-and-pull multigene strategy based on polycistronic expression and metabolic flux regulation to enhance betalain biosynthesis in transgenic maize (Zea mays L.) endosperm. We engineered a novel enhanced RUBY (eRUBY) system derived from the original polycistronic RUBY construct (CYP76AD1P2ADODA1P2ADOPA5GT unit, abbreviated CDG) by introducing arogenate dehydrogenase (ADHa) to increase the L-tyrosine substrate supply. All the genes were driven by the endosperm-specific promoter. Fusion of ADHa into a single polycistronic eRUBY construct (CDGA) produced significantly higher betanin (6.88 mg g-1 dry weight) and isobetanin (1.81 mg g-1 dry weight) levels than in CDG + A, which stacked the ADHa cassette independently with CDG. The high betalain accumulation in CDGA lines (which also exhibited higher transgene copy number) resulted in a 2.85-7.58-fold improvement in endosperm antioxidant capacity compared to WT (versus 2.48-2.80-fold in CDG + A). Importantly, transgenic plants maintained a normal phenotype. Transcriptome and metabolome analyses further indicated that metabolism of phenylalanine, alanine, aspartate, and glutamate contributes to betalain production. Hybridization with sweet corn successfully created a high-sugar eRUBY maize variety. Collectively, these results demonstrate the successful development of a novel maize germplasm with significantly enhanced nutritional value through high betalain accumulation. (c) 2025 Crop Science Society of China and Institute of Crop Science, CAAS. Production and hosting by Elsevier B.V. on behalf of KeAi Communications Co., Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Cytoplasmic male sterility (CMS) reduces pollen fertility and nuclear Rf genes restore fertility. This Commentary explores support for the "arms race" and "pre-adaptation" models for CMS-Rf co-evolution. CMS and Rf interact dynamically, influencing plant reproduction, stress tolerance, and hybrid seed production, highlighting co-evolutionary processes critical for crop improvement.

Genome editing using CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein) or other systems has become a cornerstone of numerous biological and applied research fields. However, detecting the resulting mutations by analyzing sequencing data remains time consuming and inefficient. In response to this issue, we designed SuperDecode, an integrated software toolkit for analyzing editing outcomes using a range of sequencing strategies. SuperDecode comprises three modules, DSDecodeMS, HiDecode, and LaDecode, each designed to automatically decode mutations from Sanger, high-throughput short-read, and long-read sequencing data, respectively, from targeted PCR amplicons. By leveraging specific strategies for constructing sequencing libraries of pooled multiple amplicons, HiDecode and LaDecode facilitate large-scale identification of mutations induced by single or multiplex target-site editing in a cost-effective manner. We demonstrate the efficacy of SuperDecode by analyzing mutations produced using different genome editing tools (CRISPR/Cas, base editing, and prime editing) in different materials (diploid and tetraploid rice and protoplasts), underscoring its versatility in decoding genome editing outcomes across different applications. Furthermore, this toolkit can be used to analyze other genetic variations, as exemplified by its ability to estimate the C-to-U editing rate of the cellular RNA of a mitochondrial gene. SuperDecode offers both a standalone software package and a web-based version, ensuring its easy access and broad compatibility across diverse computer systems. Thus, SuperDecode provides a comprehensive platform for analyzing a wide array of mutations, advancing the utility of genome editing for scientific research and genetic engineering.

We revealed that editing the promoter and distal regulatory region of the pleiotropic genes Ghd7 and PRR37 reduces their ability to delay heading date while improving their capacity to boost crop yield, offering valuable resources for rice breeding. Heading date is a crucial agronomic characteristic in rice that governs the adaptability to different latitudes and the yield of various varieties. Optimizing the heading date of superior cultivars in breeding practice can significantly broaden their potential planting areas. Ghd7 and PRR37 are pivotal genes that control heading date and enhance agronomic traits. In the elite indica rice variety Mei Xiang Zhan 2 (MXZ2), we used CRISPR/Cas9 technology to effectively generate homozygous mutant lines with a gradient change in heading date by multi-target editing the promoter and distal regulatory region of Ghd7 and PRR37. Various degrees of down-regulation of Ghd7 or PRR37 expression, impaired gene functions, and advancement of the heading date were observed in the mutant lines. Certain mutant lines exhibited an early heading date and increased yield while preserving the exceptional quality of MXZ2. Our study revealed that editing the promoter and distal regulatory region of the pleiotropic genes Ghd7 and PRR37 reduces their ability to delay heading date while improving their capacity to boost crop yield, offering valuable resources for rice breeding.

The canonical CRISPR/Cas editors are constrained by the requirement for specific PAMs, which substantially limits their editable target range. Rice (particularly indica rice) is sensitive to low temperature, which impacts the yield and restricts the geographic distribution of rice. In this study, we developed PAM-flexible multiplex genome editing tools based on SpG (recognising NGN-PAMs) and SpRY (recognising NNN-PAMs) variants. We then tested the feasibility of using a sweet potato leaf curl virus (SPLCV) replicon-based expression vector and single-stranded DNA-binding domain (DBD) to improve the editing efficiency of these PAM-flexible editors. Furthermore, we used SpG-mediated multiplex genome editing to achieve comprehensive improvement in cold tolerance in indica rice by editing WRKY transcription factors OsWRKY53 and OsWRKY63, to generate high cold-resistant indica rice lines. We concluded that these PAM-flexible multiplex genome editors are powerful tools for multi-gene editing for crop genetic improvement.

Soil salinization has emerged as a major threat affecting crop yields. Global warming leads to a massive loss of terrestrial water and makes soils saltier. Cultivating salt-tolerant crops is the major strategy adopted for utilizing these salinized soils. Sea Rice 86 (SR86) is one such elite salt-tolerant rice variety derived from ancient indica rice. However, SR86 has multiple wild traits, such as tallness and strong photoperiod sensitivity (PS), which have limited its application in agricultural production. In this study, we edited 13 genes responsible for 10 traits in SR86 to develop an improved SR86M line by using clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9 multiplex-genome-editing technology, high-throughput sequencing, crossing, and progeny selection. Subsequent analysis of SR86M detected nine genes with expected mutations, leading to changes in seven traits, including improvements of plant architecture, plant height and PS decreased, grain number, grain length, fragrance, and nitrogen utilization efficiency increased. The improved agronomic traits in SR86M are similar to modern cultivated rice, along with elite salt tolerance like SR86, indicating suitability for potential cultivation. Our results also reveal the efficiency of multiplex-genome-editing in directional improvement of crop varieties.

Ginsenosides, the primary bioactive components of Panax ginseng, exhibit diverse pharmacological properties, ranging from anticancer to neuroprotective effects. However, traditional production by ginseng cultivation faces limitations due to extended growth cycles, insufficient yields, intricate extraction processes, and significant environmental dependencies. Synthetic biology and synthetic metabolic engineering offer promising alternatives for sustainable manufacturing of essential bioactive compounds, including ginsenosides. First, this review describes the ginsenoside biosynthesis pathways, emphasizing crucial enzymes (e.g., HMG-CoA reductase, squalene epoxidase, dammarenediol-II synthase, amyrin synthase, and various UDP-glycosyltransferases) and their regulatory networks. Understanding these fundamental pathways enables rational engineering of production systems. Second, it examines current synthetic biology approaches, encompassing plant cell, tissue, and hairy root cultures, engineered microbial hosts including Saccharomyces cerevisiae and Escherichia coli, and cell-free enzymatic synthesis. Third, it evaluates the medicinal significance, market prospects, and industrial feasibility of these biomanufactured compounds. Finally, it analyzes the sustainability of production models and explores the emerging potential of engineered plant chassis. These advanced methodologies directly address traditional agricultural constraints and establish a robust framework for future ginsenoside synthesis.

Short tandem repeats (STRs) modulate gene expression and contribute to trait variation. However, a systematic evaluation of the genomic characteristics of STRs has not been conducted, and their influence on gene expression in rice remains unclear. Here, we construct a map of 137,629 polymorphic STRs in the rice (Oryza sativa L.) genome using a population-scale resequencing dataset. A genome-wide survey encompassing 4726 accessions shows that the occurrence frequency, mutational patterns, chromosomal distribution, and functional properties of STRs are correlated with the sequences and lengths of repeat motifs. Leveraging a transcriptome dataset from 127 rice accessions, we identify 44,672 expression STRs (eSTRs) by modeling gene expression in response to the length variation of STRs. These eSTRs are notably enriched in the regulatory regions of genes with active transcriptional signatures. Population analysis identifies numerous STRs that have undergone genetic divergence among different rice groups and 1726 tagged STRs that may be associated with agronomic traits. By editing the (ACT)7 STR in OsFD1 promoter, we further experimentally validate its role in regulating gene expression and phenotype. Our study highlights the contribution of STRs to transcriptional regulation in plants and establishes the foundation for their potential use as alternative targets for genetic improvement.

eRUBY is a powerful visual reporter combining a feedback-insensitive TyrA arogenate dehydrogenase (ADHα) gene with original RUBY structure, and substantially promotes betalain biosynthesis in rice endosperm.