Weed genomics:yielding insights into the genetics of weedy traits for crop improvement

Weeds cause tremendous economic and ecological damage worldwide.The number of genomes established for weed species has sharply increased during the recent decade,with some 26 weed species having been sequenced and de novo genomes assembled.These genomes range from 270 Mb(Barbarea vulgaris)to almost 4.4 Gb(Aegilops tauschii).Importantly,chromosome-level assemblies are now available for 17 of these 26 species,and genomic investigations on weed populations have been conducted in at least 12 species.The resulting genomic data have greatly facilitated studies of weed management and biology,especially origin and evolution.Available weed genomes have indeed revealed valuable weed-derived genetic materials for crop improvement.In this review,we summarize the recent progress made in weed genomics and provide a perspective for further exploitation in this emerging field.

Structures of plant resistosome reveal how NLR immune receptors are activated

Nucleotide-binding domain and leucine-rich repeat(NLR)proteins make up the largest immune receptor family in plants.Although many studies have put effort into revealing the working mechanism of NLRs,the activation details of plant NLRs still remain obscure.Recently,two remarkable works resolved the structures of a plant NLR protein,the Arabidopsis thaliana HOPZ-ACTIVATED RESISTANCE1(ZAR1),both in resting and activation states.The activated ZAR1 with its partner proteins form a wheel-like pentamer called resistosome that is thought to be able to trigger cell death by perturbing plasma membrane integrity.These findings greatly further our understanding of plant immune system.

Transcriptome-wide N6-methyladenosine(m^(6)A)methylation in soybean under Meloidogyne incognita infection

N^(6)-methyladenosine(m^(6)A)is a reversible epigenetic modification of mRNA and other RNAs that plays a significant role in regulating gene expression and biological processes.However,m^(6)A abundance,dynamics,and transcriptional regulatory mechanisms remain unexplored in the context of soybean resistance to Meloidogyne incognita.In this study,we performed a comparative analysis of transcriptome-wide m^(6)A and metabolome profiles of soybean root tissues with and without M.incognita infection.Global m^(6)A hypermethylation was widely induced in response to M.incognita infection and was enriched around the 3′end of coding sequences and in 3′UTR regions.There were 2069 significantly modified m^(6)A sites,594 differentially expressed genes,and 103 differentially accumulated metabolites between infected and uninfected roots,including coumestrol,psoralidin,and 2-hydroxyethylphosphonate.Among 101 m^(6)A-modified DEGs,34 genes were hypomethylated and upregulated,and 39 genes were hypermethylated and downregulated,indicating a highly negative correlation between m^(6)A methylation and gene transcript abundance.A number of these m^(6)A-modified DEGs,including WRKY70,ERF60,POD47 and LRR receptor-like serine/threonine-protein kinases,were involved in plant defense responses.Our study provides new insights into the critical role of m^(6)A modification in early soybean responses to M.incognita.

Insights into pollen-stigma recognition:self-incompatibility mechanisms serve as interspecies barriers in Brassicaceae?

A new study provides a comprehensive molecular mechanism that controls interspecific incompatibility of self-incompatible(SI)plants in the Brassicaceae.This finding points to a potentially promising path to break interspecific barriers and achieve introgression of desirable traits into crops from distant species among SI crops in the Brassicaceae.

G2-LIKE CAROTENOID REGULATOR(SlGCR)is a positive regulator of lutein biosynthesis in tomato

Lutein is an oxygen-containing carotenoid synthesized in plant chloroplasts and chromoplasts.It plays an indispensable role in promoting plant growth and maintaining eye health in humans.The rate-limiting step of lutein biosynthesis is catalyzed by the lycopeneε-cyclase enzyme(LCYE).Although great progress has been made in the identification of transcription factors involved in the lutein biosynthetic pathway,many systematic molecular mechanisms remain to be elucidated.Here,using co-expression analysis,we identified a gene,G2-LIKE CAROTENOID REGULATOR(SlGCR),encoding a GARP G2-like transcription factor,as the potential regulator of SlLCYE in tomato.Silencing of SlGCR reduced the expression of carotenoid biosynthetic genes and the accumulation of carotenoids in tomato leaves.By contrast,overexpression of SlGCR in tomato fruit significantly increased the expression of relevant genes and enhanced the accumulation of carotenoids.SlGCR can directly bind to the SlLCYE promoter and activate its expression.In addition,we also discovered that expression of SlGCR was negatively regulated by the master regulator SlRIN,thereby inhibiting lutein synthesis during tomato fruit ripening.Taken together,we identified SlGCR as a novel regulator involved in tomato lutein biosynthesis,elucidated the regulatory mechanism,and provided a potential tool for tomato lutein metabolic engineering.

Genome editing toward biofortified soybean with minimal trade-off between low phytic acid and yield

Phytic acid(PA)in grain seeds reduces the bioavailability of nutrient elements in monogastric animals,and an important objective for crop seed biofortification is to decrease the seed PA content.Here,we employed CRISPR/Cas9 to generate a PA mutant population targeting PA biosynthesis and transport genes,including two multi-drug-resistant protein 5(MRP5)and three inositol pentose-phosphate kinases(IPK1).We characterized a variety of lines containing mutations on multiple IPK and MRP5 genes.The seed PA was more significantly decreased in higher-order mutant lines with multiplex mutations.However,such mutants also exhibited poor agronomic performance.In the population,we identified two lines carrying single mutations in ipk1b and ipk1c,respectively.These mutants exhibited moderately reduced PA content,and regular agronomic performance compared to the wild type.Our study indicates that moderately decreasing PA by targeting single GmIPK1 genes,rather than multiplex mutagenesis toward ultra-low PA,is an optimal strategy for low-PA soybean with a minimal trade-off in yield performance.

From gene editing to genome engineering: restructuring plant chromosomes via CRISPR/Cas

In the last years,tremendous progress has been achieved in the field of gene editing in plants.By the induction of single site-specific double-strand breaks(DSBs),the knockout of genes by non-homologous end joining has become routine in many plant species.Recently,the efficiency of inducing pre-planned mutations by homologous recombination has also been improved considerably.However,very little effort has been undertaken until now to achieve more complex changes in plant genomes by the simultaneous induction of several DSBs.Several reports have been published on the efficient induction of deletions.However,the induction of intrachromosomal inversions and interchromosomal recombination by the use of CRISPR/Cas has only recently been reported.In this review,we want to sum up these results and put them into context with regards to what is known about natural chromosome rearrangements in plants.Moreover,we review the recent progress in CRISPR/Cas-based mammalian chromosomal rearrangements,which might be inspiring for plant biologists.In the long run,the controlled restructuring of plant genomes should enable us to link or break linkage of traits at will,thus defining a new area of plant breeding.

Coculture engineering for efficient production of vanillyl alcohol in Escherichia coli

Vanillyl alcohol is a precursor of vanillin,which is one of the most widely used flavor compounds.Currently,vanillyl alcohol biosynthesis still encounters the problem of low efficiency.In this study,coculture engineering was adopted to improve production efficiency of vanillyl alcohol in E.coli.First,two pathways were compared for biosynthesis of the immediate precursor 3,4-dihydroxybenzyl alcohol in monocultures,and the 3-dehydroshikimate-derived pathway showed higher efficiency than the 4-hydroxybenzoate-derived pathway.To enhance the efficiency of the last methylation step,two strategies were used,and strengthening S-adenosylmethionine(SAM)regeneration showed positive effect while strengthening SAM biosynthesis showed negative effect.Then,the optimized pathway was assembled in a single cell.However,the biosynthetic efficiency was still low,and was not significantly improved by modular optimization of pathway genes.Thus,coculturing engineering strategy was adopted.At the optimal inoculation ratio,the titer reached 328.9 mg/L.Further,gene aroE was knocked out to reduce cell growth and improve 3,4-DHBA biosynthesis of the upstream strain.As a result,the titer was improved to 559.4 mg/L in shake flasks and to 3.89 g/L in fed-batch fermentation.These are the highest reported titers of vanillyl alcohol so far.This work provides an effective strategy for sustainable production of vanillyl alcohol.

Metabolic engineering in woody plants: challenges, advances, and opportunities

Woody plant species represent an invaluable reserve of biochemical diversity to which metabolic engineering can be applied to satisfy the need for commodity and specialty chemicals,pharmaceuticals,and renewable energy.Woody plants are particularly promising for this application due to their low input needs,high biomass,and immeasurable ecosystem services.However,existing challenges have hindered their widespread adoption in metabolic engi neering efforts,such as long generation times,large and highly heterozygous genomes,and difficulties in transfor mation and regeneration.Recent advances in omics approaches,systems biology modeling and plant transformation and regeneration methods provide effective approaches in overcoming these outstanding challenges.Promises brought by developments in this space are steadily opening the door to widespread metabolic engineering of woody pl ants to meet the global need for a wide range of sustainably sourced chemicals and materials.

Shining in the dark:the big world of small peptides in plants

Small peptides represent a subset of dark matter in plant proteomes.Through differential expression patterns and modes of action,small peptides act as important regulators of plant growth and development.Over the past 20 years,many small peptides have been identified due to technical advances in genome sequencing,bioinformatics,and chemical biology.In this article,we summarize the classifi-cation of plant small peptides and experimental strategies used to identify them as well as their potential use in agronomic breeding.We review the biological functions and molecular mechanisms of small peptides in plants,discuss current problems in small peptide research and highlight future research directions in this field.Our review provides crucial insight into small peptides in plants and will contribute to a better understanding of their potential roles in biotechnology and agriculture.

Reprogramming plant specialized metabolism by manipulating protein kinases

Being sessile,plants have evolved sophisticated mechanisms to balance between growth and defense to survive in the harsh environment.The transition from growth to defense is commonly achieved by factors,such as protein kinases(PKs)and transcription factors,that initiate signal transduction and regulate specialized metabolism.Plants produce an array of lineage-specific specialized metabolites for chemical defense and stress tolerance.Some of these molecules are also used by humans as drugs.However,many of these defense-responsive metabolites are toxic to plant cells and inhibitory to growth and development.Plants have,thus,evolved complex regulatory networks to balance the accumulation of the toxic metabolites.Perception of external stimuli is a vital part of the regulatory network.Protein kinase-mediated signaling activates a series of defense responses by phosphorylating the target pro-teins and translating the stimulus into downstream cellular signaling.As biosynthesis of specialized metabolites is triggered when plants perceive stimuli,a possible connection between PKs and spe-cial ized meta bolism is well recognized.However,the roles of PKs in plant specialized metabolism have not received much attention until recently.Here,we summarize the recent advances in understanding PKs in plant specialized metabolism.We aim to highlight how the stimulatory signals are transduced,leading to the biosynthesis of corresponding metabolites.We discuss the post-translational regulation of specialized metabolism and provide insights into the mechanisms by which plants respond to the external signals.In addition,we propose possible strategies to increase the production of plant spe-cial ized metabolites in biotechnological applications using PKs.

Correction:Co-expression of GR79 EPSPS and GAT generates high glyphosate-resistant alfalfa with low glyphosate residues

Correction:aBIOTECH[2023]4:352-358 https://doi.org/10.1007/s42994-023-00119-3 In the Acknowledgements section of this article the funding number incorrectly given as SQ2022YF F1000033 and should have been 2022YFF1003204.

Remodeling of the cell wall as a drought-tolerance mechanism of a soybean genotype revealed by global gene expression analysis

Drought stress is major abiotic stress that affects soybean production.Therefore,it is widely desirable that soybean becomes more tolerant to stress.To provide insights into regulatory mechanisms of the stress response,we compared the global gene expression profiles from leaves of two soybean genotypes that display different responses to water-deficit(BR 16 and Embrapa 48,drought-sensitive and droughttolerant,respectively).After the RNA-seq analysis,a total of 5335 down-regulated and 3170 up-regulated genes were identified in the BR16.On the other hand,the number of genes differentially expressed was markedly lower in the Embrapa 48,355 up-regulated and 471 down-regulated genes.However,induction and expression of protein kinases and transcription factors indicated signaling cascades involved in the drought tolerance.Overall,the results suggest that the metabolism of pectin is differently modulated in response to drought stress and may play a role in the soybean defense mechanism against drought.This occurs via an increase of the cell wall plasticity and crosslink,which contributed to a higher hydraulic conductance(Kf)and relative water content(RWC%).The drought-tolerance mechanism of the Embrapa 48 genotype involves remodeling of the cell wall and increase of the hydraulic conductance to the maintenance of cell turgor and metabolic processes,resulting in the highest leaf RWC,photosynthetic rate(A),transpiration(E)and carboxylation(A/Ci).Thus,we concluded that the cell wall adjustment under drought is important for a more efficient water use which promoted a more active photosynthetic metabolism,maintaining higher plant growth under drought stress.

Multi-strategy engineering greatly enhances provitamin A carotenoid accumulation and stability in Arabidopsis seeds

Staple grains with low levels of provitamin A carotenoids contribute to the global prevalence of vitamin A deficiency and therefore are the main targets for provitamin A biofortification.However,carotenoid stability during both seed maturation and postharvest storage is a serious concern for the full benefits of carotenoid biofortified grains.In this study,we utilized Arabidopsis as a model to establish car-otenoid biofortification strategies in seeds.We discovered that manipulation of carotenoid biosynthetic activity by seed-specific expression of Phytoene synthase(PSY)increases both provitamin A and total carotenoid levels but the increased carotenoids are prone to degradation during seed maturation and storage,consistent with previous studies of provitamin A biofortified grains.In contrast,stacking with Orange(OR^(His)),a gene that initiates chromopl ast biogenesis,dramatically enhances provitamin A and total carotenoid content and stability.Up to 65-and 10-fold increases of β-carotene and total car-otenoids,res pectively,with provitamin A carotenoids composing over 63%were observed in the seeds containing OR^(His) and PSY.Co-expression of Homogen tisate geranylgeranyl transferase(HGGT)with OR^(His) and PSY further increases carotenoid accumulation and stability during seed maturation and storage.Moreover,knocking-out of B-carotene hydroxylase 2(BCH2)by CRISPR/Cas9 not only potentially facilitates β-carotene accumulation but also minimizes the negative effect of carotenoid over production on seed germi nation.Our findings provide new insights into various processes on carotenoid accu-mulation and stability in seeds and establish a multiplexed strategy to simultaneously target carotenoid biosynthesis,turnover,and stable storage for carotenoid biofortification in crop seeds.

NERD1 is required for primexine formation and plasma membrane undulation during microsporogenesis in Arabidopsis thaliana

The primexine formation and plasma membrane undulation are the crucial steps of pollen wall formation in many angiosperms.However,the molecular mechanism underlining these processes is largely unknown.In Arabidopsis,NEW ENHANCER OF ROOT DWARFISM1(NERD1),a transmembrane protein,was reported to play pleiotropic roles in plant development including male fertility control;while,how NERD1 disruption impacts male reproduction is yet unclear.Here,we revealed that the male sterility of nerd1 mutants is attributed to defects in early steps of pollen wall formation.We found that nerd1-2 is void of primexine formation and microspore plasma membrane undulation,defective in callose deposition.Consequently,sporopollenin precursors are unable to deposit and assemble on the microspore surface,but instead accumulated in the anther locule and tapetal cells,and ultimately leading to microspore abortion.NERD1 is localized in the Golgi and is expressed in both vegetative and reproductive organs,with the highest expression in reproductive tissues,including the tapetum,male meiocytes,tetrads and mature pollen grains.Our results suggest that NERD1 is required for the primexine deposition and microspore plasma membrane undulation,thus essential for sporopollenin assembly and pollen exine formation.

Single-cell profiling lights different cell trajectories in plants

The molecular mechanism of the maintenance and differentiation of plant stem cells is an eternal theme in studies on plant growth and development.Recent advances in single-cell RNA sequencing(scRNAseq)methods have completely changed the understanding of cell heterogeneity and cell function,allowing research precision to identify the differentiation trajectory of stem cells maintained and differentiated at the cellular level.This review aimed to mainly discuss the novel insights provided by scRNA-seq for the maintenance and initiation of plant stem cells,cell differentiation,cell response to environmental changes,and improvement strategies for scRNA-seq.In addition,it highlighted additional perspectives beyond scRNA-seq,such as spatial transcriptomes,epigenomes,and single-cell multiomics,for a renewed understanding of stem cell maintenance and cell differentiation,thus providing potential targets and theoretical foundations for crop improvement.

Achieving broad-spectrum resistance against rice bacterial blight through targeted promoter editing and pathogen population monitoring

Plant diseases severely reduce crop yields and threaten global food security.Broad-spectrum resistance(BSR)is a desirable trait because it confers resistance against more than one pathogen species or the majority of races/strains of the same pathogen.To control plant diseases,breeders have selected BSR to reduce disease occurrence and prolong the life-span of newly released cultivars in the last several decades(Mundt,Phytopathology 108(7):792–802,2018).Although effective,breeding of BSR cultivars in crop plants is still time-consuming and technically challenging.Recently,new gene-editing technologies such as CRISPR/Cas9 have dramatically accelerated the process of plant breeding and provided an approach for rapidly creating new varieties with BSR and other beneficial traits(Borrelli et al.,Front Plant Sci 9:1245,2018).In addition,close surveillance of pathogen populations in the field can provide useful information for the deployment of appropriate resistance genes in the target regions.In this mini-review,we focus on the significance and application of the exciting results from two recent companion papers published in Nature Biotechnology that provide new strategies to develop crop plants with BSR against pathogens through targeted promoter editing of susceptibility genes in plants as well as pathogen population monitoring.

Pepino mosaic virus antagonizes plant m^(6)A modification by promoting the autophagic degradation of the m^(6)A writer HAKAI

Autophagy plays an active anti-viral role in plants.Increasing evidence suggests that viruses can inhibit or manipulate autophagy,thereby winning the arms race between plants and viruses.Here,we demonstrate that overexpression of an m^(6)A writer from Solanum lycopersicum,SlHAKAI,could negatively regulate pepino mosaic virus(PepMV)infection,inhibit viral RNA and protein accumulations by affecting viral m^(6)A levels in tomato plants and vice versa.The PepMV-encoded RNA-dependent RNA polymerase(RdRP)directly interacts with SlHAKAI and reduces its protein accumulation.The RdRP-mediated decreased protein accumulation of SlHAKAI is sensitive to the autophagy inhibitor 3-methyladenine and is compromised by knocking down a core autophagy gene.Furthermore,PepMV RdRP could interact with an essential autophagy-related protein,SlBeclin1.RdRP,SlHAKAI,and SlBeclin1 interaction complexes form bright granules in the cytoplasm.Silencing of Beclin1 in Nicotiana benthamiana plants abolishes the RdRP-mediated degradation of SlHAKAI,indicating the requirement of Beclin1 in this process.This study uncovers that the PepMV RdRP exploits the autophagy pathway by interacting with SlBeclin1 to promote the autophagic degradation of the SlHAKAI protein,thereby inhibiting the m^(6)A modification-mediated plant defense responses.

Technological breakthroughs in generating transgene-free and genetically stable CRISPR-edited plants

CRISPR/Cas9 gene-editing technologies have been very effective in editing target genes in all major crop plants and offer unprecedented potentials in crop improvement.A major challenge in using CRISPR gene-editing technology for agricultural applications is that the target gene-edited crop plants need to be transgene free to maintain trait stability and to gain regulatory approval for commercial production.In this article,we present various strategies for generating transgene-free and target geneedited crop plants.The CRISPR transgenes can be removed by genetic segregation if the crop plants are reproduced sexually.Marker-assisted tracking and eliminating transgenes greatly decrease the time and labor needed for identifying the ideal transgene-free plants.Transgenes can be programed to undergo self-elimination when CRISPR genes and suicide genes are sequentially activated,greatly accelerating the isolation of transgene-free and target gene-edited plants.Transgene-free plants can also be generated using approaches that are considered non-transgenic such as ribonucleoprotein transfection,transient expression of transgenes without DNA integration,and nano-biotechnology.Here,we discuss the advantages and disadvantages of the various strategies in generating transgene-free plants and provide guidance for adopting the best strategies in editing a crop plant.

Molecular regulation of tomato male reproductive development

The reproductive success of flowering plants,which directly affects crop yield,is sensitive to environmental changes.A thorough understanding of how crop reproductive development adapts to climate changes is vital for ensuring global food security.In addition to being a high-value vegetable crop,tomato is also a model plant used for research on plant reproductive development.Tomato crops are cultivated under highly diverse climatic conditions worldwide.Targeted crosses of hybrid varieties have resulted in increased yields and abiotic stress resistance;however,tomato reproduction,especially male reproductive development,is sensitive to temperature fluctuations,which can lead to aborted male gametophytes,with detrimental effects on fruit set.We herein review the cytological features as well as genetic and molecular pathways influencing tomato male reproductive organ development and responses to abiotic stress.We also compare the shared features among the associated regulatory mechanisms of tomato and other plants.Collectively,this review highlights the opportunities and challenges related to characterizing and exploiting genic male sterility in tomato hybrid breeding programs.