Polyembryony has posed a significant impediment to the advancement of citrus hybrid breeding.FhRWP is widely regarded as a pivotal factor governing asexual reproduction in citrus,and prior research has demonstrated th...Polyembryony has posed a significant impediment to the advancement of citrus hybrid breeding.FhRWP is widely regarded as a pivotal factor governing asexual reproduction in citrus,and prior research has demonstrated that FhARID1,acting as an upstream regulator,modulates FhRWP expression.In this study,we performed a genome-wide characterization of the ARID-HMG-related genes using the short juvenile minicitrus Fortunella hindsii.A total of 20 ARID-HMG-related genes were identified.Protein interaction network and enrichment analysis suggested that ARID-HMG-related proteins might might be involved in chromatin remodeling complexes.Knockout of FhARID1 in F.hindsii did not induce the conversion from polyembryony to monoembryony.However,fharid1 plants in T1 generation exhibited abnormal proliferation at axillary buds,which is similar to phenotype of fhrwp plants.Expression analysis of fharid1 ovary tissues revealed the downregulation of FhRWP.The results indicated that FhARID1,as an upstream regulator of FhRWP,has an effect on the development of citrus axillary buds.Expression analysis of overexpressed leaves of FhARID1 lines showed that no significant up-regulation of FhRWP,indicating that FhARID1 is not the sole upstream regulatory factor of FhRWP.Only FhARID2 showed a correlation in expression with FhARID1 among the ARID-related genes,further supporting the notion that this gene may be involved in complex formation rather than acting alone.Yeast two-hybrid and MS/MS spectra further indicated that FhARID1 function requires casein kinase II-mediated post-transcriptional phosphorylation.This study elucidated the function of FhARID1 in citrus apomixis and axillary bud development,providing a fundamental basis for understanding the role of ARID-HMG-related genes.展开更多
Carotenoids are indispensable for both human health and plant survival.Citrus,is one of the fruit crops richest in carotenoid compounds,with approximately 115 kinds of carotenoids;tremendous diversity in carotenoids c...Carotenoids are indispensable for both human health and plant survival.Citrus,is one of the fruit crops richest in carotenoid compounds,with approximately 115 kinds of carotenoids;tremendous diversity in carotenoids composition and concentration exists among various species,showing different colors from nearly white to crimson.The carotenoid biosynthetic pathway and the key carotenogenic genes have been identified in citrus;however,the underlying regulatory mechanisms remain unclear.In this study,among the main species of genus Citrus(primitive,wild,and cultivated),we detected carotenoids in flavedo using High-Performance Liquid Chromatography,and analyzed variations in cis-acting elements in the promoters of key carotenoid pathway genes.Intriguingly,both carotenoid composition and content were generally increased during the evolution of citrus,and the corresponding variations in the promoters were identified,including the gain or loss of critical environmental stress-responsive elements and hormone-responsive elements,which are closely associated with carotenoid enhancement.In addition,pummelo has the most heat-responsive elements,but the Mangshan mandarin does not have this element in the promoters of PSY,which is highly related to their geographical origin and indicate that temperature is a critical environmental signal influencing carotenoid accumulation.Moreover,the abscisic acid-responsive motif was rich in almost all the seven species,but the ethylene-responsive motif was deficient,which demystified the unique phytohormone regulation mechanism of carotenoid accumulation in citrus.Overall,our study provides new insights into the molecular regulatory mechanism of carotenoid enhancement in the evolution of citrus,which can facilitate breeding and cultivation efforts to improve the nutritional quality and esthetic value in citrus and hopefully other fruit crops.展开更多
Carotenoids are indispensable for human health,and citrus fruit are a crucial source of dietary carotenoids.Bagging,an important orchard practice to enhance fruit economic value,is widely used in many horticultural cr...Carotenoids are indispensable for human health,and citrus fruit are a crucial source of dietary carotenoids.Bagging,an important orchard practice to enhance fruit economic value,is widely used in many horticultural crops,including citrus fruit.The bagged‘Majiayou’pomelo(Citrus grandis)produces vivid deeper red pulp,a fantastic agronomic trait,but the underlying molecular regulatory mechanism remains largely unexplored.Here,the enhancement of carotenoids,especially lycopene,was confirmed by HPLC analysis of carotenoids in the pulp of bagged fruit and controls.qRT-PCR analysis of the 14 carotenoid pathway genes further revealed that upregulated PSY and downregulated CCD1 expression in bagged fruit could directly enhance the accumulation of carotenoids.In addition,RNA-seq analysis identified 311 differentially expressed genes(DEGs)in the bagged fruit and controls in five developmental stages.Weighted gene co-expression network analysis(WGCNA)identified 13 critical candidate genes among the DEGs,which are closely associated with lycopene accumulation.The underlying regulatory mechanism of these candidate genes on the transcription of carotenoid pathway genes in the bagged fruit was discussed.Considering that the candidate genes were involved in the corresponding metabolic pathways,the increase in sucrose content and decrease in ABA in bagged fruit were also identified,implying that these candidate genes may be indirectly related to carotenoid enhancement in pulp by regulating phytohormones,primary metabolism,and stress responses.The results provide new insights into the potential regulatory mechanism of lycopene enhancement in the pulp of bagged‘Majiayou’pomelo,facilitating breeding and orchard management efforts to improve the nutritional quality and esthetic value of citrus,and perhaps other fruit crops.展开更多
Self-occlusions are common in rice canopy images and strongly influence the calculation accuracies of panicle traits. Such interference can be largely eliminated if panicles are phenotyped at the 3 D level.Research on...Self-occlusions are common in rice canopy images and strongly influence the calculation accuracies of panicle traits. Such interference can be largely eliminated if panicles are phenotyped at the 3 D level.Research on 3 D panicle phenotyping has been limited. Given that existing 3 D modeling techniques do not focus on specified parts of a target object, an efficient method for panicle modeling of large numbers of rice plants is lacking. This paper presents an automatic and nondestructive method for 3 D panicle modeling. The proposed method integrates shoot rice reconstruction with shape from silhouette, 2 D panicle segmentation with a deep convolutional neural network, and 3 D panicle segmentation with ray tracing and supervoxel clustering. A multiview imaging system was built to acquire image sequences of rice canopies with an efficiency of approximately 4 min per rice plant. The execution time of panicle modeling per rice plant using 90 images was approximately 26 min. The outputs of the algorithm for a single rice plant are a shoot rice model, surface shoot rice model, panicle model, and surface panicle model, all represented by a list of spatial coordinates. The efficiency and performance were evaluated and compared with the classical structure-from-motion algorithm. The results demonstrated that the proposed method is well qualified to recover the 3 D shapes of rice panicles from multiview images and is readily adaptable to rice plants of diverse accessions and growth stages. The proposed algorithm is superior to the structure-from-motion method in terms of texture preservation and computational efficiency. The sample images and implementation of the algorithm are available online. This automatic, cost-efficient, and nondestructive method of 3 D panicle modeling may be applied to high-throughput 3 D phenotyping of large rice populations.展开更多
Carotenoids directly influence citrus fruit color and nutritional value,which is critical to consumer acceptance.Elucidating the potential molecular mechanism underlying carotenoid metabolism is of great importance fo...Carotenoids directly influence citrus fruit color and nutritional value,which is critical to consumer acceptance.Elucidating the potential molecular mechanism underlying carotenoid metabolism is of great importance for improving fruit quality.Despite the well-established carotenoid biosynthetic pathways,the molecular regulatory mechanism underlying carotenoid metabolism remains poorly understood.Our previous studies have reported that the Myc-type basic helix-loop-helix(bHLH)transcription factor(TF)regulates citrus proanthocyanidin biosynthesis.Transgenic analyses further showed that overexpression of CsTT8 could significantly promote carotenoid accumulation in transgenic citrus calli,but its regulatory mechanism is still unclear.In the present study,we found that overexpression of CsTT8 enhances carotenoid content in citrus fruit and calli by increasing the expression of CsDXR,CsHDS,CsHDR,CsPDS,CsLCYE,CsZEP,and CsNCED2,which was accompanied by changes in the contents of abscisic acid and gibberellin.The in vitro and in vivo assays indicated that CsTT8 directly bound to the promoters of CsDXR,CsHDS,and CsHDR,the keymetabolic enzymes of the methylerythritol 4-phosphate(MEP)pathway,thus providing precursors for carotenoid biosynthesis and transcriptionally activating the expression of these three genes.In addition,CsTT8 activated the promoters of four key carotenoid biosynthesis pathway genes,CsPDS,CsLCYE,CsZEP,and CsNCED2,directly promoting carotenoid biosynthesis.This study reveals a novel network of carotenoid metabolism regulated by CsTT8.Our findings will contribute to manipulating carotenoid metabolic engineering to improve the quality of citrus fruit and other crops.展开更多
Self-incompatibility(SI)substantially restricts the yield and quality of citrus.Therefore,breeding and analyzing selfcompatible germplasm is of great theoretical and practical signi ficance for citrus.Here,we focus on...Self-incompatibility(SI)substantially restricts the yield and quality of citrus.Therefore,breeding and analyzing selfcompatible germplasm is of great theoretical and practical signi ficance for citrus.Here,we focus on the mechanism of a self-compatibility mutation in‘Guiyou No.1'pummelo(Citrus maxima),which is a spontaneous mutant of‘Shatian’pummelo(Citrus maxima,self-incompatibility).The rate of fruit set and the growth of pollen tubes in the pistil con firmed that a spontaneous mutation in the pistil is responsible for the self-compatibility of‘Guiyou No.1'.Segregation ratios of the S genotype in progeny,expression analysis,and western blotting validated that the reduced levels of S_(2)-RNase mRNA contribute to the loss of SI in‘Guiyou No.1'.Furthermore,we report a phased assembly of the‘Guiyou No.1'pummelo genome and obtained two complete and well-annotated S haplotypes.Coupled with an analysis of SV variations,methylation levels,and gene expression,we identi fied a candidate gene(CgHB40),that may in fluence the regulation of the S/^RNase promoter.Our data provide evidence that a mutation that affects the pistilled to the loss of SI in‘Guiyou No.1'by in fluencing a poorly understood mechanism that affects transcriptional regulation.This work signi ficantly advances our understanding of the genetic basis of the SI system in citrus and provides information on the regulation of S-RNase genes.展开更多
Deciphering the genetic basis of plant secondary metabolism will provide useful insights for genetic improvement and enhance our fundamental understanding of plant biological processes.Although citrus plants are among...Deciphering the genetic basis of plant secondary metabolism will provide useful insights for genetic improvement and enhance our fundamental understanding of plant biological processes.Although citrus plants are among the most important fruit crops worldwide,the genetic basis of secondary metabolism in these plants is largely unknown.Here,we use a high-density linkage map to dissect large-scale flavonoid metabolic traits measured in different tissues(young leaf,old leaf,mature pericarp,and mature pulp)of an F_(1) pseudo-testcross citrus population.We detected 80 flavonoids in this population and identified 138 quantitative trait loci(QTLs)for 57 flavonoids in these four tissues.Based on transcriptional profiling and functional annotation,twenty-one candidate genes were identified,and one gene encoding flavanone 3-hydroxylase(F3H)was functionally verified to result in naturally occurring variation in dihydrokaempferol content through genetic variations in its promoter and coding regions.The abundant data resources collected for diverse citrus germplasms here lay the foundation for complete characterization of the citrus flavonoid biosynthetic pathway and will thereby promote efficient utilization of metabolites in citrus quality improvement.展开更多
Elaioplasts of citrus peel are colorless plastids which accumulate significant amounts of terpenes.However,other functions of elaioplasts have not been fully characterized to date.Here,a LC–MS/MS shotgun technology w...Elaioplasts of citrus peel are colorless plastids which accumulate significant amounts of terpenes.However,other functions of elaioplasts have not been fully characterized to date.Here,a LC–MS/MS shotgun technology was applied to identify the proteins from elaioplasts that were highly purified from young fruit peel of kumquat.A total of 655 putative plastid proteins were identified from elaioplasts according to sequence homology in silico and manual curation.Based on functional classification via Mapman,~50%of the identified proteins fall into six categories,including protein metabolism,transport,and lipid metabolism.Of note,elaioplasts contained ATP synthase and ADP,ATP carrier proteins at high abundance,indicating important roles for ATP generation and transport in elaioplast biogenesis.Additionally,a comparison of proteins between citrus chromoplast and elaioplast proteomes suggest a high level of functional conservation.However,some distinctive protein profiles were also observed in both types of plastids notably for isoprene biosynthesis in elaioplasts,and carotenoid metabolism in chromoplasts.In conclusion,this comprehensive proteomic study provides new insights into the major metabolic pathways and unique characteristics of elaioplasts and chromoplasts in citrus fruit.展开更多
Abstract As an essential horticultural crop,Citrus has carotenoid diversity,which affects its aesthetic and nutritional values.β,β-Xanthophylls are the primary carotenoids accumulated in citrus fruits,and non-heme d...Abstract As an essential horticultural crop,Citrus has carotenoid diversity,which affects its aesthetic and nutritional values.β,β-Xanthophylls are the primary carotenoids accumulated in citrus fruits,and non-heme di-iron carotene hydroxylase(BCH)enzymes are mainly responsible forβ,β-xanthophyll synthesis.Previous studies have focused on the hydroxylation of BCH1,but the role of its paralogous gene in citrus,BCH2,remains largely unknown.In this study,we revealed theβ-hydroxylation activity of citrus BCH2(CsBCH2)for the first time through the functional complementation assay using Escherichia coli,although CsBCH2 exhibited a lower activity in hydroxylatingβ-carotene intoβ-cryptoxanthin than citrus BCH1(CsBCH1).Our results showed that overexpression of CsBCH2 in citrus callus increased xanthophyll proportion and plastoglobule size with feedback regulation of carotenogenic gene expression.This study revealed the distinct expression patterns and functional characteristics of two paralogous genes,CsBCH1 and CsBCH2,and illustrated the backup compensatory role of CsBCH2 for CsBCH1 in citrus xanthophyll biosynthesis.The independent function of CsBCH2 and its cooperative function with CsBCH1 inβ-cryptoxanthin biosynthesis suggested the potential of CsBCH2 to be employed for expanding the synthetic biology toolkit in carotenoid engineering。展开更多
Ethylene treatment promotes orange coloration in the flavedo of Satsuma mandarin(Citrus unshiu Marc.)fruit,but the corresponding regulatory mechanism is still largely unknown.In this study,we identified a C2H2-type zi...Ethylene treatment promotes orange coloration in the flavedo of Satsuma mandarin(Citrus unshiu Marc.)fruit,but the corresponding regulatory mechanism is still largely unknown.In this study,we identified a C2H2-type zinc-finger transcription factor,CitZAT4,the expression of which was markedly induced by ethylene.CitZAT4 directly binds to the Cit PSY promoter and activates its expression,thereby promoting carotenoid biosynthesis.Transient expression in Satsuma mandarin fruit and stable transformation of citrus calli showed that overexpressing of CitZAT4 inhibited Cit LCYE expression,thus inhibitingα-branch yellow carotenoid(lutein)biosynthesis.CitZAT4 overexpression also enhanced the transcript levels of CitLCYB,Cit HYD,and CitNCED2,promotingβ-branch orange carotenoid accumulation.Molecular biochemical assays,including yeast one-hybrid(Y1H),electrophoretic mobility shift(EMSA),chromatin immunoprecipitation quantitative polymerase chain reaction(Ch IP-q PCR),and luciferase(LUC)assays,demonstrated that CitZAT4 directly binds to the promoters of its target genes and regulates their expression.An ethylene response factor,CitERF061,which is induced by ethylene signaling,was found to directly bound to the CitZAT4 promoter and induced its expression,thus positively regulating CitZAT4-mediated orange coloration in citrus fruit.Together,our findings reveal that a CitZAT4-mediated transcriptional cascade is driven by ethylene via CitERF061,linking ethylene signaling to carotenoid metabolism in promoting orange coloration in the flavedo of Satsuma mandarin fruit.The molecular regulatory mechanism revealed here represents a significant step toward developing strategies for improving the quality and economic efficiency of citrus crops.展开更多
Carotenoids and apocarotenoids act as phytohormones and volatile precursors that influence plant development and confer aesthetic and nutritional value critical to consumer preference.Citrus fruits display considerabl...Carotenoids and apocarotenoids act as phytohormones and volatile precursors that influence plant development and confer aesthetic and nutritional value critical to consumer preference.Citrus fruits display considerable natural variation in carotenoid and apocarotenoid pigments.In this study,using an integrated genetic approach we revealed that a 5;c/s-regulatory change at CCD4b encoding CAROTENOID CLEAVAGE DIOXYGENASE 4b is a major genetic determinant of natural variation in C3 0 apocarotenoids responsible for red coloration of citrus peel.Functional analyses demonstrated that in addition the known role in synthesizing 3-citraurin,CCD4b is also responsible for the production of another important C3 0 apocarotenoid pigment,p-citraurinene.Furthermore,analyses of the CCD4b promoter and transcripts from various citrus germplasm accessions established a tight correlation between the presence of a putative 5'c/s-regulatory enhancer within an MITE transposon and the enhanced allelic expression of CCD4b in C3 0 apocarotenoid-rich red-peeled accessions.Phylogenetic analysis provided further evidence that functional diversification of CCD4b and naturally occurring variation of the CCD4b promoter resulted in the stepwise evolution of red peels in mandarins and their hybrids.Taken together,our findings provide new insights into the genetic and evolutionary basis of apocarotenoid diversity in plants,and would facilitate breeding efforts that aim to improve the nutritional and aesthetic value of citrus and perhaps other fruit crops.展开更多
Carotenoid biosynthesis is closely associated with abscisic acid(ABA)during the ripening process of non-climacteric fruits,but the regulatory mechanism that links ABA signaling to carotenoid metabolism remains largely...Carotenoid biosynthesis is closely associated with abscisic acid(ABA)during the ripening process of non-climacteric fruits,but the regulatory mechanism that links ABA signaling to carotenoid metabolism remains largely unclear.Here,we identified two master regulators of ABA-mediated citrus fruit coloration,CsERF110 and CsERF53,which activate the expression of carotenoid metabolism genes(CsGGPPS,CsPSY,CsPDS,CsCRTISO,CsLCYB2,CsLCYE,CsHYD,CsZEP,and CsNCED2)to facilitate carotenoid accumulation.Further investigations showed that CsERF110 not only activates the expression of CsERF53 by binding to its promoter but also interacts with CsERF53 to form the transcriptional regulatory module CsERF110-CsERF53.We also discovered a positive feedback regulatory loop between the ABA signal and carotenoid metabolism regulated by the transcriptional regulatory module CsERF110-CsERF53.Our results reveal that the CsERF110-CsERF53 module responds to ABA signaling,thereby orchestrating citrus fruit coloration.Considering the importance of carotenoid content for citrus and many other carotenoid-rich crops,the revelation of molecular mechanisms that underlie ABA-mediated carotenoid biosynthesis in plants will facilitate the development of transgenic/gene-editing approaches,further contributing to improving the quality of citrus and other carotenoid-rich crops.展开更多
Carotenoids in plant foods provide health benefits by functioning as provitamin A.One ofthe vital provitamin A carotenoids,β-cryptoxanthin,is typically plentiful in citrus fruit.However,little is known about the gene...Carotenoids in plant foods provide health benefits by functioning as provitamin A.One ofthe vital provitamin A carotenoids,β-cryptoxanthin,is typically plentiful in citrus fruit.However,little is known about the genetic basis of β-cryptoxanthin accumulation in citrus.Here,we performed a widely targeted metabolomic analysis of 65 major carotenoids and carotenoid derivatives to characterize carotenoid accumulation in Citrus and determine the taxonomic profile of b-cryptoxanthin.We used data from 81 newly sequenced representative accessions and 69 previously sequenced Citrus cultivars to reveal the genetic basis of β-cryptoxanthin accumulation through a genome-wide association study.We identified a causal gene,CitCYP97B,which encodes a cytochrome P450 protein whose substrate and metabolic pathways in land plants were undetermined.We subsequently demonstrated that CitCYP97B functions as a novel monooxygenase that specifically hydroxylates the β-ring of β-cryptoxanthin in a heterologous expression system.In planta experiments provided further evidence that CitCYP97B negatively regulates b-cryptoxanthin content.Using the sequenced Citrus accessions,we found that two critical structural cis-element variations contribute to increased expression of CitCYP97B,thereby altering β-cryptoxanthin accumulation in fruit.Hybridization/introgression appear to have contributed to the prevalence of two cis-element variations in different Citrus types during citrus evolution.Overall,these findings extend our understanding of the regulation and diversity of carotenoid metabolism in fruit crops and provide a genetic target for production of β-cryptoxanthin-biofortified products.展开更多
Kumquat(Fortunella spp.)is a fruit and ornamental crop worldwide due to the palatable taste and high ornamental value of its fruit.Although Fortunella is classified into the economically important true citrus fruit tr...Kumquat(Fortunella spp.)is a fruit and ornamental crop worldwide due to the palatable taste and high ornamental value of its fruit.Although Fortunella is classified into the economically important true citrus fruit tree group together with Citrus and Poncirus,few studies have been focused on its evolutionary scenario.In this study,analysis of five chloroplast loci and 47 nuclear microsatellites(nSSR)loci from 38 kumquat and 10 citrus accessions revealed the independent phylogeny of Fortunella among citrus taxa,and that Fortunella mainly comprises two populations:CUL,cultivated Fortunella spp.(F.margarita,F.crassifolia and F.japonica);and HK,wild Hong Kong kumquat(Fortunella hindsii).Genomic analysis based on whole-genome SNPs indicated that the allele frequency of both pupations deviated from the neutral selection model,suggesting directional selection was a force driving their evolutions.CUL exhibited lower genomic diversity and higher linkage strength than HK,suggesting artificial selection involved in its origin.A high level of genetic differentiation(Fst=0.364)was detected and obviously asynchronous demographic changes were observed between CUL and HK.Based on these results,a new hypothesis for the speciation of Fortunella is proposed.展开更多
Dear Editor,In trans-specific S-haplotypes,crossover recombination at the S-locus,which controls self-incompatibility,has been thought to be highly suppressed.Here,we discovered a previously unreported super S haploty...Dear Editor,In trans-specific S-haplotypes,crossover recombination at the S-locus,which controls self-incompatibility,has been thought to be highly suppressed.Here,we discovered a previously unreported super S haplotype,derived from a rare inter-haplotypic recombination event,that contained two complete suites of functional S-RNase and SLF genes and could break the SI response in trifoliate orange through“self-recognition”within the same pollen.展开更多
Verticillium wilt is one of the most critical cotton diseases,which is widely distributed in cotton-producing countries.However,the conventional method of verticillium wilt investigation is still manual,which has the ...Verticillium wilt is one of the most critical cotton diseases,which is widely distributed in cotton-producing countries.However,the conventional method of verticillium wilt investigation is still manual,which has the disadvantages of subjectivity and low efficiency.In this research,an intelligent vision-based system was proposed to dynamically observe cotton verticillium wilt with high accuracy and high throughput.Firstly,a 3-coordinate motion platform was designed with the movement range 6,100 mm×950 mm×500 mm,and a specific control unit was adopted to achieve accurate movement and automatic imaging.Secondly,the verticillium wilt recognition was established based on 6 deep learning models,in which the VarifocalNet(VFNet)model had the best performance with a mean average precision(mAP)of 0.932.Meanwhile,deformable convolution,deformable region of interest pooling,and soft non-maximum suppression optimization methods were adopted to improve VFNet,and the mAP of the VFNet-Improved model improved by 1.8%.The precision–recall curves showed that VFNet-Improved was superior to VFNet for each category and had a better improvement effect on the ill leaf category than fine leaf.The regression results showed that the system measurement based on VFNet-Improved achieved high consistency with manual measurements.Finally,the user software was designed based on VFNet-Improved,and the dynamic observation results proved that this system was able to accurately investigate cotton verticillium wilt and quantify the prevalence rate of different resistant varieties.In conclusion,this study has demonstrated a novel intelligent system for the dynamic observation of cotton verticillium wilt on the seedbed,which provides a feasible and effective tool for cotton breeding and disease resistance research.展开更多
基金funded by the National Key Research and Development Program of China(Grant No.2022YFF1003100)Modern Citrus Industry Technology System of China(Grant No.CARS-26).
文摘Polyembryony has posed a significant impediment to the advancement of citrus hybrid breeding.FhRWP is widely regarded as a pivotal factor governing asexual reproduction in citrus,and prior research has demonstrated that FhARID1,acting as an upstream regulator,modulates FhRWP expression.In this study,we performed a genome-wide characterization of the ARID-HMG-related genes using the short juvenile minicitrus Fortunella hindsii.A total of 20 ARID-HMG-related genes were identified.Protein interaction network and enrichment analysis suggested that ARID-HMG-related proteins might might be involved in chromatin remodeling complexes.Knockout of FhARID1 in F.hindsii did not induce the conversion from polyembryony to monoembryony.However,fharid1 plants in T1 generation exhibited abnormal proliferation at axillary buds,which is similar to phenotype of fhrwp plants.Expression analysis of fharid1 ovary tissues revealed the downregulation of FhRWP.The results indicated that FhARID1,as an upstream regulator of FhRWP,has an effect on the development of citrus axillary buds.Expression analysis of overexpressed leaves of FhARID1 lines showed that no significant up-regulation of FhRWP,indicating that FhARID1 is not the sole upstream regulatory factor of FhRWP.Only FhARID2 showed a correlation in expression with FhARID1 among the ARID-related genes,further supporting the notion that this gene may be involved in complex formation rather than acting alone.Yeast two-hybrid and MS/MS spectra further indicated that FhARID1 function requires casein kinase II-mediated post-transcriptional phosphorylation.This study elucidated the function of FhARID1 in citrus apomixis and axillary bud development,providing a fundamental basis for understanding the role of ARID-HMG-related genes.
基金This research was supported by National Key Research and Development Program of China(Grant No.2018YFD1000200)National Natural Science Foundation of China(Grant nos.31930095 and 31630065)We should thank Prof.Zuoxiong Liu for editing the English language of the manuscript.
文摘Carotenoids are indispensable for both human health and plant survival.Citrus,is one of the fruit crops richest in carotenoid compounds,with approximately 115 kinds of carotenoids;tremendous diversity in carotenoids composition and concentration exists among various species,showing different colors from nearly white to crimson.The carotenoid biosynthetic pathway and the key carotenogenic genes have been identified in citrus;however,the underlying regulatory mechanisms remain unclear.In this study,among the main species of genus Citrus(primitive,wild,and cultivated),we detected carotenoids in flavedo using High-Performance Liquid Chromatography,and analyzed variations in cis-acting elements in the promoters of key carotenoid pathway genes.Intriguingly,both carotenoid composition and content were generally increased during the evolution of citrus,and the corresponding variations in the promoters were identified,including the gain or loss of critical environmental stress-responsive elements and hormone-responsive elements,which are closely associated with carotenoid enhancement.In addition,pummelo has the most heat-responsive elements,but the Mangshan mandarin does not have this element in the promoters of PSY,which is highly related to their geographical origin and indicate that temperature is a critical environmental signal influencing carotenoid accumulation.Moreover,the abscisic acid-responsive motif was rich in almost all the seven species,but the ethylene-responsive motif was deficient,which demystified the unique phytohormone regulation mechanism of carotenoid accumulation in citrus.Overall,our study provides new insights into the molecular regulatory mechanism of carotenoid enhancement in the evolution of citrus,which can facilitate breeding and cultivation efforts to improve the nutritional quality and esthetic value in citrus and hopefully other fruit crops.
基金supported by the National Key Research and Development Program of China (Grant No. 2018YFD1000200)National Natural Science Foundation of China (Grant No. 31930095, 31630065)
文摘Carotenoids are indispensable for human health,and citrus fruit are a crucial source of dietary carotenoids.Bagging,an important orchard practice to enhance fruit economic value,is widely used in many horticultural crops,including citrus fruit.The bagged‘Majiayou’pomelo(Citrus grandis)produces vivid deeper red pulp,a fantastic agronomic trait,but the underlying molecular regulatory mechanism remains largely unexplored.Here,the enhancement of carotenoids,especially lycopene,was confirmed by HPLC analysis of carotenoids in the pulp of bagged fruit and controls.qRT-PCR analysis of the 14 carotenoid pathway genes further revealed that upregulated PSY and downregulated CCD1 expression in bagged fruit could directly enhance the accumulation of carotenoids.In addition,RNA-seq analysis identified 311 differentially expressed genes(DEGs)in the bagged fruit and controls in five developmental stages.Weighted gene co-expression network analysis(WGCNA)identified 13 critical candidate genes among the DEGs,which are closely associated with lycopene accumulation.The underlying regulatory mechanism of these candidate genes on the transcription of carotenoid pathway genes in the bagged fruit was discussed.Considering that the candidate genes were involved in the corresponding metabolic pathways,the increase in sucrose content and decrease in ABA in bagged fruit were also identified,implying that these candidate genes may be indirectly related to carotenoid enhancement in pulp by regulating phytohormones,primary metabolism,and stress responses.The results provide new insights into the potential regulatory mechanism of lycopene enhancement in the pulp of bagged‘Majiayou’pomelo,facilitating breeding and orchard management efforts to improve the nutritional quality and esthetic value of citrus,and perhaps other fruit crops.
基金supported by the National Natural Science Foundation of China (U21A20205)Key Projects of Natural Science Foundation of Hubei Province (2021CFA059)+1 种基金Fundamental Research Funds for the Central Universities (2021ZKPY006)cooperative funding between Huazhong Agricultural University and Shenzhen Institute of Agricultural Genomics (SZYJY2021005,SZYJY2021007)。
文摘Self-occlusions are common in rice canopy images and strongly influence the calculation accuracies of panicle traits. Such interference can be largely eliminated if panicles are phenotyped at the 3 D level.Research on 3 D panicle phenotyping has been limited. Given that existing 3 D modeling techniques do not focus on specified parts of a target object, an efficient method for panicle modeling of large numbers of rice plants is lacking. This paper presents an automatic and nondestructive method for 3 D panicle modeling. The proposed method integrates shoot rice reconstruction with shape from silhouette, 2 D panicle segmentation with a deep convolutional neural network, and 3 D panicle segmentation with ray tracing and supervoxel clustering. A multiview imaging system was built to acquire image sequences of rice canopies with an efficiency of approximately 4 min per rice plant. The execution time of panicle modeling per rice plant using 90 images was approximately 26 min. The outputs of the algorithm for a single rice plant are a shoot rice model, surface shoot rice model, panicle model, and surface panicle model, all represented by a list of spatial coordinates. The efficiency and performance were evaluated and compared with the classical structure-from-motion algorithm. The results demonstrated that the proposed method is well qualified to recover the 3 D shapes of rice panicles from multiview images and is readily adaptable to rice plants of diverse accessions and growth stages. The proposed algorithm is superior to the structure-from-motion method in terms of texture preservation and computational efficiency. The sample images and implementation of the algorithm are available online. This automatic, cost-efficient, and nondestructive method of 3 D panicle modeling may be applied to high-throughput 3 D phenotyping of large rice populations.
基金supported by the National Natural Science Foundation of China(No.31930095,32172527)the Modern Agro-industry Technology Research System(CARS-26).
文摘Carotenoids directly influence citrus fruit color and nutritional value,which is critical to consumer acceptance.Elucidating the potential molecular mechanism underlying carotenoid metabolism is of great importance for improving fruit quality.Despite the well-established carotenoid biosynthetic pathways,the molecular regulatory mechanism underlying carotenoid metabolism remains poorly understood.Our previous studies have reported that the Myc-type basic helix-loop-helix(bHLH)transcription factor(TF)regulates citrus proanthocyanidin biosynthesis.Transgenic analyses further showed that overexpression of CsTT8 could significantly promote carotenoid accumulation in transgenic citrus calli,but its regulatory mechanism is still unclear.In the present study,we found that overexpression of CsTT8 enhances carotenoid content in citrus fruit and calli by increasing the expression of CsDXR,CsHDS,CsHDR,CsPDS,CsLCYE,CsZEP,and CsNCED2,which was accompanied by changes in the contents of abscisic acid and gibberellin.The in vitro and in vivo assays indicated that CsTT8 directly bound to the promoters of CsDXR,CsHDS,and CsHDR,the keymetabolic enzymes of the methylerythritol 4-phosphate(MEP)pathway,thus providing precursors for carotenoid biosynthesis and transcriptionally activating the expression of these three genes.In addition,CsTT8 activated the promoters of four key carotenoid biosynthesis pathway genes,CsPDS,CsLCYE,CsZEP,and CsNCED2,directly promoting carotenoid biosynthesis.This study reveals a novel network of carotenoid metabolism regulated by CsTT8.Our findings will contribute to manipulating carotenoid metabolic engineering to improve the quality of citrus fruit and other crops.
基金This research was financially supported by the National Key Research and Development Program of China(grant no.2018YFD1000107)the National Natural Science Foundation of China(grant nos.31772259,31630065,and 31521092)+1 种基金the Fundamental Research Funds forthe Central Univer sities(grant no.2662019PY044)the China Agriculture Research System of MOF and MARA and the Hubei Provincial Natural Science Foundation of China(2020CFB532).
文摘Self-incompatibility(SI)substantially restricts the yield and quality of citrus.Therefore,breeding and analyzing selfcompatible germplasm is of great theoretical and practical signi ficance for citrus.Here,we focus on the mechanism of a self-compatibility mutation in‘Guiyou No.1'pummelo(Citrus maxima),which is a spontaneous mutant of‘Shatian’pummelo(Citrus maxima,self-incompatibility).The rate of fruit set and the growth of pollen tubes in the pistil con firmed that a spontaneous mutation in the pistil is responsible for the self-compatibility of‘Guiyou No.1'.Segregation ratios of the S genotype in progeny,expression analysis,and western blotting validated that the reduced levels of S_(2)-RNase mRNA contribute to the loss of SI in‘Guiyou No.1'.Furthermore,we report a phased assembly of the‘Guiyou No.1'pummelo genome and obtained two complete and well-annotated S haplotypes.Coupled with an analysis of SV variations,methylation levels,and gene expression,we identi fied a candidate gene(CgHB40),that may in fluence the regulation of the S/^RNase promoter.Our data provide evidence that a mutation that affects the pistilled to the loss of SI in‘Guiyou No.1'by in fluencing a poorly understood mechanism that affects transcriptional regulation.This work signi ficantly advances our understanding of the genetic basis of the SI system in citrus and provides information on the regulation of S-RNase genes.
文摘Deciphering the genetic basis of plant secondary metabolism will provide useful insights for genetic improvement and enhance our fundamental understanding of plant biological processes.Although citrus plants are among the most important fruit crops worldwide,the genetic basis of secondary metabolism in these plants is largely unknown.Here,we use a high-density linkage map to dissect large-scale flavonoid metabolic traits measured in different tissues(young leaf,old leaf,mature pericarp,and mature pulp)of an F_(1) pseudo-testcross citrus population.We detected 80 flavonoids in this population and identified 138 quantitative trait loci(QTLs)for 57 flavonoids in these four tissues.Based on transcriptional profiling and functional annotation,twenty-one candidate genes were identified,and one gene encoding flavanone 3-hydroxylase(F3H)was functionally verified to result in naturally occurring variation in dihydrokaempferol content through genetic variations in its promoter and coding regions.The abundant data resources collected for diverse citrus germplasms here lay the foundation for complete characterization of the citrus flavonoid biosynthetic pathway and will thereby promote efficient utilization of metabolites in citrus quality improvement.
基金This work was supported by the National Natural Science Foundation of China(NSFC,grant nos.31501739)the National Basic Research Program of China(973 project No.2013CB127105)Huazhong Agricultural University Scientific&Technological Self-innovation Foundation(No.2662015BQ034).
文摘Elaioplasts of citrus peel are colorless plastids which accumulate significant amounts of terpenes.However,other functions of elaioplasts have not been fully characterized to date.Here,a LC–MS/MS shotgun technology was applied to identify the proteins from elaioplasts that were highly purified from young fruit peel of kumquat.A total of 655 putative plastid proteins were identified from elaioplasts according to sequence homology in silico and manual curation.Based on functional classification via Mapman,~50%of the identified proteins fall into six categories,including protein metabolism,transport,and lipid metabolism.Of note,elaioplasts contained ATP synthase and ADP,ATP carrier proteins at high abundance,indicating important roles for ATP generation and transport in elaioplast biogenesis.Additionally,a comparison of proteins between citrus chromoplast and elaioplast proteomes suggest a high level of functional conservation.However,some distinctive protein profiles were also observed in both types of plastids notably for isoprene biosynthesis in elaioplasts,and carotenoid metabolism in chromoplasts.In conclusion,this comprehensive proteomic study provides new insights into the major metabolic pathways and unique characteristics of elaioplasts and chromoplasts in citrus fruit.
基金supported by the National Natural Science Foundation of China(No.31930095 and 32172527)the Modern Agro-industry Technology Research System(CARS-26).
文摘Abstract As an essential horticultural crop,Citrus has carotenoid diversity,which affects its aesthetic and nutritional values.β,β-Xanthophylls are the primary carotenoids accumulated in citrus fruits,and non-heme di-iron carotene hydroxylase(BCH)enzymes are mainly responsible forβ,β-xanthophyll synthesis.Previous studies have focused on the hydroxylation of BCH1,but the role of its paralogous gene in citrus,BCH2,remains largely unknown.In this study,we revealed theβ-hydroxylation activity of citrus BCH2(CsBCH2)for the first time through the functional complementation assay using Escherichia coli,although CsBCH2 exhibited a lower activity in hydroxylatingβ-carotene intoβ-cryptoxanthin than citrus BCH1(CsBCH1).Our results showed that overexpression of CsBCH2 in citrus callus increased xanthophyll proportion and plastoglobule size with feedback regulation of carotenogenic gene expression.This study revealed the distinct expression patterns and functional characteristics of two paralogous genes,CsBCH1 and CsBCH2,and illustrated the backup compensatory role of CsBCH2 for CsBCH1 in citrus xanthophyll biosynthesis.The independent function of CsBCH2 and its cooperative function with CsBCH1 inβ-cryptoxanthin biosynthesis suggested the potential of CsBCH2 to be employed for expanding the synthetic biology toolkit in carotenoid engineering。
基金supported by the National Key R&D Program of China(2023YFD2300600)the National Natural Science Foundation of China(No.31930095)the National Modern Agricultural(Citrus)Technology Systems of China(No.CARS-27)。
文摘Ethylene treatment promotes orange coloration in the flavedo of Satsuma mandarin(Citrus unshiu Marc.)fruit,but the corresponding regulatory mechanism is still largely unknown.In this study,we identified a C2H2-type zinc-finger transcription factor,CitZAT4,the expression of which was markedly induced by ethylene.CitZAT4 directly binds to the Cit PSY promoter and activates its expression,thereby promoting carotenoid biosynthesis.Transient expression in Satsuma mandarin fruit and stable transformation of citrus calli showed that overexpressing of CitZAT4 inhibited Cit LCYE expression,thus inhibitingα-branch yellow carotenoid(lutein)biosynthesis.CitZAT4 overexpression also enhanced the transcript levels of CitLCYB,Cit HYD,and CitNCED2,promotingβ-branch orange carotenoid accumulation.Molecular biochemical assays,including yeast one-hybrid(Y1H),electrophoretic mobility shift(EMSA),chromatin immunoprecipitation quantitative polymerase chain reaction(Ch IP-q PCR),and luciferase(LUC)assays,demonstrated that CitZAT4 directly binds to the promoters of its target genes and regulates their expression.An ethylene response factor,CitERF061,which is induced by ethylene signaling,was found to directly bound to the CitZAT4 promoter and induced its expression,thus positively regulating CitZAT4-mediated orange coloration in citrus fruit.Together,our findings reveal that a CitZAT4-mediated transcriptional cascade is driven by ethylene via CitERF061,linking ethylene signaling to carotenoid metabolism in promoting orange coloration in the flavedo of Satsuma mandarin fruit.The molecular regulatory mechanism revealed here represents a significant step toward developing strategies for improving the quality and economic efficiency of citrus crops.
基金the National Key R&D Program of China(2018YFD1000200)the National Natural Science Foundation of China(no.31630065,31330066 and 31521092)China Agriculture Research System(CARS-27)the 111 project(B13034),and a China Council Scholarship(to X.Z.).
文摘Carotenoids and apocarotenoids act as phytohormones and volatile precursors that influence plant development and confer aesthetic and nutritional value critical to consumer preference.Citrus fruits display considerable natural variation in carotenoid and apocarotenoid pigments.In this study,using an integrated genetic approach we revealed that a 5;c/s-regulatory change at CCD4b encoding CAROTENOID CLEAVAGE DIOXYGENASE 4b is a major genetic determinant of natural variation in C3 0 apocarotenoids responsible for red coloration of citrus peel.Functional analyses demonstrated that in addition the known role in synthesizing 3-citraurin,CCD4b is also responsible for the production of another important C3 0 apocarotenoid pigment,p-citraurinene.Furthermore,analyses of the CCD4b promoter and transcripts from various citrus germplasm accessions established a tight correlation between the presence of a putative 5'c/s-regulatory enhancer within an MITE transposon and the enhanced allelic expression of CCD4b in C3 0 apocarotenoid-rich red-peeled accessions.Phylogenetic analysis provided further evidence that functional diversification of CCD4b and naturally occurring variation of the CCD4b promoter resulted in the stepwise evolution of red peels in mandarins and their hybrids.Taken together,our findings provide new insights into the genetic and evolutionary basis of apocarotenoid diversity in plants,and would facilitate breeding efforts that aim to improve the nutritional and aesthetic value of citrus and perhaps other fruit crops.
基金National Key R&D Program of China(2023YFD2300600)National Natural Science Foundation of China(no.31930095)National Modern Agricultural(Citrus)Technology Systems of China(no.CARS-27).
文摘Carotenoid biosynthesis is closely associated with abscisic acid(ABA)during the ripening process of non-climacteric fruits,but the regulatory mechanism that links ABA signaling to carotenoid metabolism remains largely unclear.Here,we identified two master regulators of ABA-mediated citrus fruit coloration,CsERF110 and CsERF53,which activate the expression of carotenoid metabolism genes(CsGGPPS,CsPSY,CsPDS,CsCRTISO,CsLCYB2,CsLCYE,CsHYD,CsZEP,and CsNCED2)to facilitate carotenoid accumulation.Further investigations showed that CsERF110 not only activates the expression of CsERF53 by binding to its promoter but also interacts with CsERF53 to form the transcriptional regulatory module CsERF110-CsERF53.We also discovered a positive feedback regulatory loop between the ABA signal and carotenoid metabolism regulated by the transcriptional regulatory module CsERF110-CsERF53.Our results reveal that the CsERF110-CsERF53 module responds to ABA signaling,thereby orchestrating citrus fruit coloration.Considering the importance of carotenoid content for citrus and many other carotenoid-rich crops,the revelation of molecular mechanisms that underlie ABA-mediated carotenoid biosynthesis in plants will facilitate the development of transgenic/gene-editing approaches,further contributing to improving the quality of citrus and other carotenoid-rich crops.
基金supported by the National Key Research and Develop-ment Program of China(2022YFF1003100)the National Natural Science Foundation of China(31930095)Modern Agro-industry Technology Research System(CARS-26).
文摘Carotenoids in plant foods provide health benefits by functioning as provitamin A.One ofthe vital provitamin A carotenoids,β-cryptoxanthin,is typically plentiful in citrus fruit.However,little is known about the genetic basis of β-cryptoxanthin accumulation in citrus.Here,we performed a widely targeted metabolomic analysis of 65 major carotenoids and carotenoid derivatives to characterize carotenoid accumulation in Citrus and determine the taxonomic profile of b-cryptoxanthin.We used data from 81 newly sequenced representative accessions and 69 previously sequenced Citrus cultivars to reveal the genetic basis of β-cryptoxanthin accumulation through a genome-wide association study.We identified a causal gene,CitCYP97B,which encodes a cytochrome P450 protein whose substrate and metabolic pathways in land plants were undetermined.We subsequently demonstrated that CitCYP97B functions as a novel monooxygenase that specifically hydroxylates the β-ring of β-cryptoxanthin in a heterologous expression system.In planta experiments provided further evidence that CitCYP97B negatively regulates b-cryptoxanthin content.Using the sequenced Citrus accessions,we found that two critical structural cis-element variations contribute to increased expression of CitCYP97B,thereby altering β-cryptoxanthin accumulation in fruit.Hybridization/introgression appear to have contributed to the prevalence of two cis-element variations in different Citrus types during citrus evolution.Overall,these findings extend our understanding of the regulation and diversity of carotenoid metabolism in fruit crops and provide a genetic target for production of β-cryptoxanthin-biofortified products.
基金funded by the National Key Research and Development Program of China(2018YFD1000106)the National Natural Science Foundation of China(31630065)Special Project for External Science and Technology Cooperation of Science and Technology Department of Yunnan Province(202003AD150014)。
文摘Kumquat(Fortunella spp.)is a fruit and ornamental crop worldwide due to the palatable taste and high ornamental value of its fruit.Although Fortunella is classified into the economically important true citrus fruit tree group together with Citrus and Poncirus,few studies have been focused on its evolutionary scenario.In this study,analysis of five chloroplast loci and 47 nuclear microsatellites(nSSR)loci from 38 kumquat and 10 citrus accessions revealed the independent phylogeny of Fortunella among citrus taxa,and that Fortunella mainly comprises two populations:CUL,cultivated Fortunella spp.(F.margarita,F.crassifolia and F.japonica);and HK,wild Hong Kong kumquat(Fortunella hindsii).Genomic analysis based on whole-genome SNPs indicated that the allele frequency of both pupations deviated from the neutral selection model,suggesting directional selection was a force driving their evolutions.CUL exhibited lower genomic diversity and higher linkage strength than HK,suggesting artificial selection involved in its origin.A high level of genetic differentiation(Fst=0.364)was detected and obviously asynchronous demographic changes were observed between CUL and HK.Based on these results,a new hypothesis for the speciation of Fortunella is proposed.
基金supported by the National Natural Science Foundation of China(grants 32122075,32302489,and 32072523)the Hubei Provincial Natural Science Foundation of China(grant 2023AFB094)+1 种基金the China Agricultural Research System(grant CARS-26)supported by the Biotechnology and Biological Sciences Research Council(BBSRC)grant BB/T00486X/1 to M.B.and Vernonica E.Franklin-Tong.
文摘Dear Editor,In trans-specific S-haplotypes,crossover recombination at the S-locus,which controls self-incompatibility,has been thought to be highly suppressed.Here,we discovered a previously unreported super S haplotype,derived from a rare inter-haplotypic recombination event,that contained two complete suites of functional S-RNase and SLF genes and could break the SI response in trifoliate orange through“self-recognition”within the same pollen.
基金supported by grants from the Major Project of Hubei Hongshan Laboratory(2022hszd004)the National Natural Science Foundation of China(32270431 and U21A20205)+1 种基金the Key Research and Development Plan of Hubei Province(2022BBA0045 and 2020000071)the Fundamental Research Funds for the Central Universities(2662022YJ018 and 2662019QD053).
文摘Verticillium wilt is one of the most critical cotton diseases,which is widely distributed in cotton-producing countries.However,the conventional method of verticillium wilt investigation is still manual,which has the disadvantages of subjectivity and low efficiency.In this research,an intelligent vision-based system was proposed to dynamically observe cotton verticillium wilt with high accuracy and high throughput.Firstly,a 3-coordinate motion platform was designed with the movement range 6,100 mm×950 mm×500 mm,and a specific control unit was adopted to achieve accurate movement and automatic imaging.Secondly,the verticillium wilt recognition was established based on 6 deep learning models,in which the VarifocalNet(VFNet)model had the best performance with a mean average precision(mAP)of 0.932.Meanwhile,deformable convolution,deformable region of interest pooling,and soft non-maximum suppression optimization methods were adopted to improve VFNet,and the mAP of the VFNet-Improved model improved by 1.8%.The precision–recall curves showed that VFNet-Improved was superior to VFNet for each category and had a better improvement effect on the ill leaf category than fine leaf.The regression results showed that the system measurement based on VFNet-Improved achieved high consistency with manual measurements.Finally,the user software was designed based on VFNet-Improved,and the dynamic observation results proved that this system was able to accurately investigate cotton verticillium wilt and quantify the prevalence rate of different resistant varieties.In conclusion,this study has demonstrated a novel intelligent system for the dynamic observation of cotton verticillium wilt on the seedbed,which provides a feasible and effective tool for cotton breeding and disease resistance research.
