This commentary spotlights landmark work from Guo et al. resolving the structure of the octameric resistosome formed by the active CCG10-nucleotide-binding leucine-rich repeat (NLR) immune receptor WAI3, which triggers s...This commentary spotlights landmark work from Guo et al. resolving the structure of the octameric resistosome formed by the active CCG10-nucleotide-binding leucine-rich repeat (NLR) immune receptor WAI3, which triggers sustained, multi-phasic Ca influx. This fills a major knowledge gap regarding EDVID-lacking plant NLRs and provides insight into plant effector-triggered immunity.
Histone demethylases have been extensively characterized in plants, yet how their enzymatic activities are regulated remains largely unknown. Here, we identify that MBD8, a protein harboring a putative methyl-CpG-binding...Histone demethylases have been extensively characterized in plants, yet how their enzymatic activities are regulated remains largely unknown. Here, we identify that MBD8, a protein harboring a putative methyl-CpG-binding domain (MBD), forms a protein complex with the histone demethylase LDL2 in Arabidopsis thaliana. In this complex, MBD8 functions as a cofactor that is required for efficient LDL2-mediated removal of mono- and di-methylation at histone H3 (H3K4me1 and H3K4me2) across the genome. In the Arabidopsis C24 ecotype, loss-of-function mutations in MBD8 and LDL2 result in indistinguishable late-flowering phenotypes, which are consistent with increased enrichment of H3K4me1 and H3K4me2 at the flowering repressor loci MAF1 and MAF5. Importantly, we demonstrate that the rice (Oryza sativa) MBD8 orthologs MBD708, MBD709, and MBD715 also interact with the LDL2 ortholog Hd18 to form a protein complex, and disruption of these components delays flowering by reprogramming the expression of genes associated with floral transition. Collectively, this study reveals an evolutionarily conserved mechanism in which MBD8 orthologs act as cofactors of LDL2 orthologs to regulate histone demethylation and flowering time in plants.
Gene editing of three key genes, G PROTEIN GAMMA SUBUNIT, CYTOKININ OXIDASE, AND IDEAL PLANT ARCHITECTURE 1, improved the salt-tolerant japonica rice variety Lansheng increased yield while maintaining its original salt t...Gene editing of three key genes, G PROTEIN GAMMA SUBUNIT, CYTOKININ OXIDASE, AND IDEAL PLANT ARCHITECTURE 1, improved the salt-tolerant japonica rice variety Lansheng increased yield while maintaining its original salt tolerance and quality.
Thermogenesis represents one of the oldest pollination traits; yet, the ecological function of the microclimates that it creates remains poorly understood. Here, we elucidate how the spatiotemporally dynamic microclimate...Thermogenesis represents one of the oldest pollination traits; yet, the ecological function of the microclimates that it creates remains poorly understood. Here, we elucidate how the spatiotemporally dynamic microclimate of Alocasia odora orchestrates pollinator movement within its inflorescence. Upregulation of alternative oxidase (AOX) in the upper spadix generates a steep, diurnally fluctuating thermal gradient against the non-thermogenic lower chamber, forming a "thermal map". We demonstrate that this thermal push-pull mechanism, interacting synergistically with the insects' innate requirements, drives the cyclical vertical movements of its specialized Colocasiomyia pollinators. While strict feeding requirements establish a baseline preference for the lower sterile zone, species-specific oviposition preferences dictate their distinct movement trajectories. Specifically, C. xenalocasiae primarily drives pollen importation, whereas C. alocasiae acts as the dominant contributor to pollen exportation. In contrast, floral scent primarily functions in long-distance pollinator attraction, while light plays only a minor modulatory role in this fine-scale within-inflorescence navigation. Our comparative analysis reveals a prevalent spatiotemporal separation of sexual organs across thermogenic flowers globally. We therefore propose that microclimate-guided pollinator movement-where thermal gradients overcome structural constraints to ensure reproductive success-constitutes a widespread, yet underappreciated evolutionary strategy.
Because agriculture is confronted with escalating climatic and resource challenges, next-generation breeding requires innovative strategies to sustain and enhance crop productivity. As the foundation of carbon fixation i...Because agriculture is confronted with escalating climatic and resource challenges, next-generation breeding requires innovative strategies to sustain and enhance crop productivity. As the foundation of carbon fixation in plants, photosynthesis strongly affects crop yield potential. Therefore, improving photosynthetic performance remains a central goal for crop improvement. Plants show substantial natural genetic variation in photosynthetic traits, arising from heritable differences in physiology, including metabolism and regulation, which represent valuable genetic resources for crop breeding. Meanwhile, advances in synthetic biology and photosynthetic genetic engineering provide complementary avenues for enhancing photosynthetic capacity and productivity. In this review, we analyze and synthesize recent progress in research on (i) natural variation in photosynthetic traits across physiological, developmental, and canopy scales; (ii) molecular and genetic regulatory mechanisms underlying photosynthetic diversity and adaptations; (iii) links between photosynthetic efficiency, source-sink coordination, and yield formation; and (iv) emerging strategies for engineering photosynthesis. We also outline remaining challenges and future perspectives. Collectively, these insights provide a strategic framework for leveraging natural genetic diversity and modern biotechnologies to optimize photosynthesis, enhance yield potential, and improve crop resilience under future climate scenarios, further supporting global food security.
Sugarcane Breeding Array 1.0, a high-density single-nucleotide polymorphism array, is based on deep whole-genome resequencing of diverse sugarcane accessions. Validation demonstrated its robust performance and cost-effec...Sugarcane Breeding Array 1.0, a high-density single-nucleotide polymorphism array, is based on deep whole-genome resequencing of diverse sugarcane accessions. Validation demonstrated its robust performance and cost-effectiveness in population structure analysis, genome-wide association studies, and molecular breeding applications.
Wang B, Wang QY, Shi WJ
… +16 more, Li RH, Zhang XD, Wang LP, Xia SJ, Chen KX, Zhou JJ, Sun YY, He J, Fernie AR, Han YC, Li JZ, Shi YH, Liu XQ, Zhang YY, Luo J, Jin C
Vitamin B6 (VB6) is an essential nutrient that must be obtained from the diet by humans and livestock, and plays an important role in response to various biotic and abiotic stresses in plants. Although the VB6 biosynthes...Vitamin B6 (VB6) is an essential nutrient that must be obtained from the diet by humans and livestock, and plays an important role in response to various biotic and abiotic stresses in plants. Although the VB6 biosynthesis pathway has been uncovered in higher plants, understanding of its genetic control remains limited, and the role of VB6 in responding to cold stress is largely unknown. Here, we showed that variation in VB6 levels in rice is attributable to differential transcription of OsPDX1.2, a pyridoxal phosphate synthase, due to polymorphisms in its promoter. Natural variations in the OsPDX1.2 promoter affect the binding affinity of OsbZIP18, a transcriptional factor repressing OsPDX1.2 transcription. Loss of function in OsPDX1.2 or overexpression of OsbZIP18 significantly reduces cold tolerance in rice. Further investigation revealed that both genes affect cold tolerance by regulating VB6 and abscisic acid (ABA) biosynthesis. Moreover, the frequency of the cold-tolerant pOsPDX1.2 increases at higher latitudes, whereas the cold-sensitive pOsPDX1.2 increases at lower latitudes, indicating that natural variation in OsPDX1.2 contributes to differential cold adaptation in rice. Overall, our results reveal a regulatory pathway controlling rice VB6 accumulation and cold tolerance via the OsbZIP18-OsPDX1.2 module, providing a potential target for improving rice quality and stress tolerance.
Meiotic crossover (CO) exchanges genetic information between homologs, thereby promoting genetic diversity among offspring. COs are non-randomly distributed across chromosomes, tending to occur in euchromatin, but rarely...Meiotic crossover (CO) exchanges genetic information between homologs, thereby promoting genetic diversity among offspring. COs are non-randomly distributed across chromosomes, tending to occur in euchromatin, but rarely in heterochromatin. In plants, H3 lysine 27 monomethylation (H3K27me1) is crucial for maintaining heterochromatin condensation and genome stability in somatic cells; however, its role in germline cells remains to be determined. Here, we demonstrate that the plant-specific H3K27 mono-methyltransferases ATXR5/6 (ARABIDOPSIS TRITHORAX-RELATED PROTEIN 5/6) play an important role in inhibiting CO formation in meiotic heterochromatin. In atxr5 atxr6, both ZMM-dependent Type I COs and ZMM-independent Type II COs are significantly increased. We further observed decondensation, decreased H3K27me1 signals, and specifically compromised non-CG methylation in atxr5 atxr6 meiotic heterochromatin. Unexpectedly, in contrast to their roles in somatic cells, where ATXR5/6 primarily regulate heterochromatin condensation and gene silencing without influencing DNA methylation, in meiocytes, ATXR5/6 mainly function in suppressing recombination and preserving heterochromatic DNA methylation without directly regulating gene expression. Moreover, loss of Type II CO regulator MMS AND UV SENSITIVE 81 (MUS81) leads to pericentromeric fragmentation and polyad formation during meiosis in the absence of ATXR5/6, indicating that MUS81 is critical for resolving atypical recombination intermediates in pericentromeric heterochromatin. Taken together, our results provide insights into the roles of ATXR5/6 in repressing meiotic recombination within heterochromatin by regulating chromosome compaction and modifications.
Auxin, primarily indole-3-acetic acid (IAA), is a central regulator of growth and development in land plants, but its physiological role in chlorophyte algae remains unclear. Here, we show that exogenous IAA modulates gr...Auxin, primarily indole-3-acetic acid (IAA), is a central regulator of growth and development in land plants, but its physiological role in chlorophyte algae remains unclear. Here, we show that exogenous IAA modulates growth in Chlorella sorokiniana, Chlorella variabilis, and Chlamydomonas reinhardtii in a concentration-dependent manner. Low IAA concentrations promoted growth by accelerating the onset of cell division without affecting cell size, whereas higher concentrations inhibited proliferation. Radiotracer assays showed that all three species take up and release IAA across the plasma membrane through a combination of passive diffusion and energy-dependent, saturable processes. Competition by excess unlabeled natural and synthetic auxins further supported the presence of carrier-mediated transport with broad substrate recognition. Phylogenetic analyses identified potential PIN-like auxin exporters in chlorophytes and other non-plant eukaryotes, and structural modeling supported conservation of the overall PIN fold and predicted auxin-binding residues. However, functional assays in Xenopus laevis oocytes, tobacco BY-2 cultured cells, and Arabidopsis thaliana did not support a role for these proteins in directional auxin export. Instead, non-plant PIN homologs localized predominantly to the endoplasmic reticulum and showed limited or no transport activity in heterologous systems. Together, these findings indicate that auxin responsiveness and basic cellular auxin transport predate canonical PIN-mediated directional auxin export, which appears to be a later innovation of the streptophyte lineage.
Despite achieving substantial progress in caloric food security, micronutrient deficiencies remain a major global health challenge, highlighting a persistent disconnect between crop productivity and nutritional quality....Despite achieving substantial progress in caloric food security, micronutrient deficiencies remain a major global health challenge, highlighting a persistent disconnect between crop productivity and nutritional quality. Crop biofortification has emerged as a promising strategy to address this gap by enhancing the intrinsic nutrient content of widely consumed foods to meet dietary requirements through complementary roles of staple crops and horticultural species in improving diet quality. In this review, we reposition biofortification within a broader nutrition-sensitive food system framework and discuss two complementary approaches to accelerate its impact. First, we examine how systematic exploration of global crop diversity can identify nutritionally superior germplasm from seed/genebanks for deployment in breeding and food systems. Second, we evaluate advances in modern breeding and genome-editing approaches for improving minerals, vitamins, and health-promoting phytochemicals across major crop groups. We further propose that a conserved set of nutrient-regulatory pathways provides a unifying framework for cross-crop biofortification, enabling more efficient, scalable strategies to enhance multiple nutritional traits to support a transition toward crop portfolios designed not only for yield and resilience but also for improved human health in the 21st century.
Inducing adventitious buds from the base of watermelon cotyledon through direct organogenesis resulted in a watermelon genetic transformation system that overcomes regeneration barriers and genotype limitations.Inducing adventitious buds from the base of watermelon cotyledon through direct organogenesis resulted in a watermelon genetic transformation system that overcomes regeneration barriers and genotype limitations.
Flowering time is a key agronomic trait that influences plant reproductive success and crop yield, and its regulation is closely associated with soil nutrient availability. This review summarizes recent advances in under...Flowering time is a key agronomic trait that influences plant reproductive success and crop yield, and its regulation is closely associated with soil nutrient availability. This review summarizes recent advances in understanding the molecular mechanisms through which macronutrients and micronutrients regulate flowering time in plants. An emerging central theme is that nutrient-derived signaling integrates with core flowering regulatory pathways, including the photoperiodic, gibberellin, vernalization, autonomous, and sugar pathways, with nitrogen and phosphorus being the most extensively studied. However, major knowledge gaps remain regarding the regulatory roles of potassium, sulfur, and micronutrients, as well as species-specific nutrient responses and the molecular basis of nutrient-nutrient and nutrient-environment interactions in flowering regulation. In addition, the role of nutrient-derived metabolites and rhizosphere microorganisms in flowering control remains largely unexplored. Addressing these challenges is essential for the rational development of crop varieties with optimized flowering time and enhanced nutrient use efficiency through targeted genetic engineering, molecular breeding, and innovative nutrient management strategies, thereby supporting sustainable agricultural development.
Yang T, Liu N, Huang L
… +17 more, Guo J, Xue X, Wang M, Zhang X, Quan C, Feng E, Tan J, Zhang Z, You Y, Xia Y, Huai D, Chen Y, Yan L, Liu K, Jiang H, Lei Y, Liao B
Peanut (Arachis hypogaea) is an allotetraploid legume crop of agricultural and economic importance worldwide. Despite advances in peanut genomics, the integration of genetic diversity with multi-omics landscapes remains...Peanut (Arachis hypogaea) is an allotetraploid legume crop of agricultural and economic importance worldwide. Despite advances in peanut genomics, the integration of genetic diversity with multi-omics landscapes remains limited, and a comprehensive understanding of subgenome asymmetry is still lacking. Here, we present a high-quality genome assembly of an elite peanut cultivar Zhonghua 5 (ZH05; 2,622.85 Mb with 73,718 predicted genes) and construct a species-specific pan-genome based on 1,286 accessions, uncovering 849.82 Mb of non-reference sequences and 18,484 novel genes. Over 23 million genetic variants and extensive gene PAVs substantially expand the landscape of genetic diversity in peanut. Integrative multi-omics (transcriptome, DNA methylation, chromatin accessibility, and three-dimensional genome architecture), combined with population genomics analyses, reveal pronounced and multi-layered subgenome asymmetry. Specifically, SubA shows higher chromatin accessibility and overall gene expression, whereas SubB displays more transposable elements, elevated DNA methylation, more genes under selection during domestication, and stronger three-dimensional chromatin interactions with more stable topologically associating domains. Furthermore, genetic dissection using a recombinant inbred line population identified several major stable QTLs for seed size, whose pyramiding contributes to ZH05's superior yield performance. Collectively, this study establishes an integrated genomic framework for peanut, illuminates asymmetric subgenome evolution across sequence, epigenetic, and chromatin architecture, and provides valuable resources for accelerating molecular breeding and crop improvement.
Loquat (Eriobotrya japonica Lindl.) is a valuable subtropical fruit tree whose genetic improvement has been significantly constrained by the absence of an efficient genetic transformation system. Although Agrobacterium-m...Loquat (Eriobotrya japonica Lindl.) is a valuable subtropical fruit tree whose genetic improvement has been significantly constrained by the absence of an efficient genetic transformation system. Although Agrobacterium-mediated transformation is the most widely used method, it proves ineffective in loquat due to the species' recalcitrance to in vitro regeneration. Pollen-based transformation offers a promising alternative by bypassing the need for tissue culture. However, the pollen wall poses a major physical barrier to the uptake of exogenous DNA. In this study, we investigated magnetic nanoparticle (MNP)-mediated transformation as a novel strategy for loquat. We confirmed that loquat pollen contains tricolporate apertures with diameters ranging from 3.0 to 5.0 μm, which are structurally suitable for the entry of MNPs-DNA. Based on this finding, we developed and optimized a transformation protocol using polyethyleneimine-coated FeO nanoparticles to deliver genetic material into loquat pollen grains. Using this approach, we successfully generated stable transgenic loquat lines, including both overexpression and gene-edited mutants. To our knowledge, this is the first report of successful MNP-mediated pollen transformation in a woody plant species. This work establishes a robust and efficient genetic transformation platform for loquat, providing a valuable tool for functional genomics and molecular breeding, as well as a potentially applicable strategy for other recalcitrant woody plants.
Co-editing the broomcorn millet PmBADH2a and PmBADH2b genes using CRISPR/Cas12i.3 generated double mutants with significantly increased 2-acetyl-1-pyrroline content, producing fragrant broomcorn millet without compromisi...Co-editing the broomcorn millet PmBADH2a and PmBADH2b genes using CRISPR/Cas12i.3 generated double mutants with significantly increased 2-acetyl-1-pyrroline content, producing fragrant broomcorn millet without compromising major agronomic traits.
Identifying male-sterile genes and developing biotechnology-based male-sterility systems are crucial for advancing hybrid maize breeding. However, this progress is hampered by the limited number of characterized key regu...Identifying male-sterile genes and developing biotechnology-based male-sterility systems are crucial for advancing hybrid maize breeding. However, this progress is hampered by the limited number of characterized key regulators and their incomplete mechanistic understanding in male sterility, as well as fertility instability of male-sterile lines and a lack of efficient maintainer lines in maize. Here, we elucidate the multifaceted roles of ZmMS1, an LBD transcription repressor. ZmMS1 coordinates timely tapetal PCD by repressing ROS-scavenging genes to regulate anther ROS homeostasis, while its DNA-binding activity is redox-sensitive, suggesting a potential redox-dependent feedback mechanism. In addition, ZmMS1 balances lipid allocation between anther cuticle and pollen exine by directly repressing sporopollenin biosynthesis and indirectly promoting cutin/wax formation. Constitutive overexpression of ZmMs1 induces dwarfism associated with GA and ABA homeostasis, and produces ~50% sterile and small pollen grains in maize and rice, offering a potential route for dwarf and male-sterile breeding. Leveraging the findings that loss and precocious expression of ZmMs1 cause recessive and dominant sterility, respectively, we develop a non-transgenic multi-control sterility system and a dominant genic male-sterility system, both showing stable and complete sterility across diverse backgrounds without yield penalty, thereby providing flexible options for hybrid maize breeding. Our findings reveal that ZmMS1, as a redox-sensitive transcription factor, regulates male fertility with previously unrevealed mechanisms, and provide practical tools for efficient hybrid maize breeding.
Mitogen-activated protein kinase (MAPK/MPK) cascades are conserved signaling modules that play crucial roles in plant growth, reproduction, and responses to biotic and abiotic stresses. Many breakthroughs have been made...Mitogen-activated protein kinase (MAPK/MPK) cascades are conserved signaling modules that play crucial roles in plant growth, reproduction, and responses to biotic and abiotic stresses. Many breakthroughs have been made recently in understanding the functional and regulatory mechanisms involving MAPK cascades in plant immunity. In this review, we summarize the conserved composition and activation of MAPK cascades and update advances in understanding the molecular mechanisms by which plants maintain or inhibit MAPK activation. We also highlight the functional links between MAPK activation and other immune events, suggesting crosstalk in the plant immune signaling network. Furthermore, we outline the strategies by which pathogen effectors inhibit plant immunity by manipulating MAPK signaling and illuminate the contribution of MAPK cascades to effector-triggered immunity. Overall, this review provides major evidence for understanding the importance and complexity of MAPK signaling and reveals that the MAPK cascade serves as an essential signaling hub in activating plant immunity.
Tissue culture recalcitrance limits plant transformation and genome editing for crop improvement. The development of new, generic factors that enhance plant regeneration efficiency could alleviate this bottleneck. Throug...Tissue culture recalcitrance limits plant transformation and genome editing for crop improvement. The development of new, generic factors that enhance plant regeneration efficiency could alleviate this bottleneck. Through precursor gene overexpression, exogenous peptide application, and loss-of-function studies, we show here that CLAVATA3/ESR-RELATED 41 (CLE41), CLE42, CLE44, and CLE46, collectively referred to as TDIF-related genes, function redundantly to promote de novo shoot regeneration. CLE41, CLE42, and CLE44 are abundantly and dynamically expressed in pluripotent callus, whereas CLE46 is shoot-induction medium-activated and specifically expressed at explant wound sites. TDIF-related peptides act through PHLOEM INTERCALATED WITH XYLEM/TDIF RECEPTOR (PXY/TDR)-subfamily members to activate WUSCHEL (WUS)-RELATED HOMEOBOX 14 (WOX14) and enhance shooting capacity. In addition, we find that LATERAL ORGAN BOUNDARIES DOMAIN4 (LBD4) is highly expressed throughout the callus and its expression increases upon TDIF upregulation in a PXY/PXL-dependent manner. Consistently, LBD4 overexpression notably improves regeneration efficiency, whereas its loss decreases regeneration. TDIF-enhanced shoot regeneration is significantly suppressed in lbd4 mutants, indicating that LBD4 is required for TDIF signaling in shoot regeneration enhancement. Furthermore, WOX14 directly activates LBD4 to promote de novo shoot regeneration. Exogenous application of synthetic TDIF-related peptides substantially increases shooting capacity in the recalcitrant crop species Sorghum bicolor L., offering a convenient new avenue to enhance de novo shoot regeneration in crops. In conclusion, we demonstrate that Arabidopsis TDIF-PXY/PXL ligand-receptor pairs markedly boost de novo shoot regeneration through WOX14-potentiated LBD4 induction. Our findings elucidate the molecular basis for a novel function of TDIF-related peptides and their receptors in hormone-driven shoot regeneration and demonstrate their practical application in improving adventitious shoot regeneration in a recalcitrant plant species.
Rising global temperature threatens fruit quality. As one of the most economically valuable fruits worldwide, citrus suffers from high-temperature-induced chlorophyll retention during ripening, significantly reducing com...Rising global temperature threatens fruit quality. As one of the most economically valuable fruits worldwide, citrus suffers from high-temperature-induced chlorophyll retention during ripening, significantly reducing commercial value. Comparative physiological analyses revealed that citrus fruit stored at 30°C exhibited markedly impaired chlorophyll degradation and failed to undergo normal degreening, in stark contrast to those maintained at 20°C. This temperature-dependent suppression of degreening was closely associated with a significant downregulation of key chlorophyll catabolic genes (CCGs). By integrating weighted gene co-expression network analysis (WGCNA) of a stay-green mutant, we identified the transcription factor CsbHLH60, a nuclear-localized transcriptional activator, from a chlorophyll-associated module. CsbHLH60 directly binds E-box motifs in the promoters of CsSGR and CsRCCR to drive their expression, a finding validated by ChIP-qPCR and EMSA assays. Transient overexpression of CsbHLH60 in citrus peel accelerated chlorophyll degradation and, importantly, overcame high-temperature-induced repression to restore degreening, whereas RNAi-mediated silencing at 20°C significantly impeded normal degreening of the peel. Mechanistically, high temperature imposes a dual "double-lock" suppression on CsbHLH60, concurrently repressing its transcription and promoting its protein degradation, thereby depleting functional activator levels. Collectively, our findings establish CsbHLH60 as a pivotal temperature-responsive regulator that directly couples temperature signaling to chlorophyll catabolism and is capable of overriding the high-temperature-induced suppression of chlorophyll degradation, thereby highlighting its potential as a molecular target for mitigating climate-driven declines in citrus fruit appearance quality.
Enhancing crop tolerance to multiple abiotic stresses is critical for achieving sustainable agriculture. Targeted seed-stage interventions using natural signaling compounds (e.g., melatonin) provide a unique opportunity...Enhancing crop tolerance to multiple abiotic stresses is critical for achieving sustainable agriculture. Targeted seed-stage interventions using natural signaling compounds (e.g., melatonin) provide a unique opportunity to establish early stress tolerance that can persist through the critical seed-to-seedling transition. Melatonin seed priming (MSP) is rapidly emerging as a green and climate-smart strategy for enhancing plant stress tolerance. MSP triggers defensive molecular, biochemical, and physiological reprogramming during germination, thereby improving plant performance under subsequent stress conditions. This review synthesizes recent mechanistic insights into how MSP confers stress tolerance across diverse species by modulating redox signaling, hormonal homeostasis, and stress-related gene networks. We elucidate the synergistic potential of MSP when combined with nanoformulations, other priming agents, or beneficial microbes. We also discuss its crosstalk with key signaling pathways to better understand the tolerance mechanisms. Furthermore, we propose a forward-looking strategy that integrates omics, genome editing, speed breeding, and molecular phenotyping methods to improve MSP applications for the development of stress-smart crops. Despite its potential, MSP still faces multiple challenges, including species-specific responses, dosage variability, limited post-priming seed storage stability, and a lack of field-scale validation. Addressing these bottlenecks through high-throughput screening, epigenetic memory assessment, and optimized delivery systems will be essential to fully harness the practical potential of MSP as a sustainable and green approach for future agriculture.