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Control of ovule development in vitis vinifera by VvMADS28 and interacting genes

Horticulture Research 2023年第6期10卷 43-55页

作者： Songlin Zhang Li Wang Jin Yao Na Wu Bilal Ahmad Steve van Nocker Jiuyun Wu Riziwangguli Abudureheman Zhi Li Xiping Wang State Key Laboratory of Crop Stress Biology in Arid Areas College of HorticultureNorthwest A&F UniversityYanglingShaanxi 712100China Key Laboratory of Horticultural Plant Biology and Germplasm Innovation in Northwest China Ministry of AgricultureNorthwest A&F UniversityYanglingShaanxi 712100China College of Horticulture Hebei Agricultural UniversityBaoding 071000China Agriculture Genomics Institute Chinese Academy of Agricultural SciencesShenzhen 518000China Department of Horticulture Michigan State UniversityEast LansingMI 48823USA Turpan Research Institute of Agricultural Sciences Xinjiang Academy of Agricultural SciencesTurpan 838000XinjiangChina

Seedless grapes are increasingly popular throughout the world,and the development of seedless varieties is a major breeding goal.In this study,we demonstrate an essential role for the grapevine MADS-box gene VvMADS28 in morphogenesis of the ovule.We found that VvMADS28 mRNA accumulated in the ovules of a seeded cultivar,‘Red Globe’,throughout the course of ovule and seed development,especially within the integument/seed coat.In contrast,in the seedless cultivar‘Thompson Seedless’,VvMADS28 was expressed only weakly in ovules,and this was associated with increased levels of histone H3 lysine 27 trimethylation(H3K27me3)within the VvMADS28 promoter region.RNAi-mediated transient suppression of VvMADS28 expression in‘Red Globe’led to reduced seed size associated with inhibition of episperm and endosperm cell development.Heterologous overexpression of VvMADS28 in transgenic tomatoes interfered with sepal development and resulted in smaller fruit but did not obviously affect seed size.Assays in yeast cells showed that VvMADS28 is subject to regulation by the transcription factor VvERF98,and that VvMADS28 could interact with the Type I/MβMADS-domain protein VvMADS5.Moreover,through DNA-affinity purification-sequencing(DAP-seq),we found that VvMADS28 protein specifically binds to the promoter of the grapevine WUSCHEL(VvWUS)gene,suggesting that maintenance of the VvMADS28–VvMADS5 dimer and VvWUS expression homeostasis influences seed development.Taken together,our results provide insight into regulatory mechanisms of ovule and seed development associated with VvMADS28.

关键词： vitis homeostasis breeding

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Discovery and genome-guided mapping of REN12 from vitis amurensis, conferring strong, rapid resistance to grapevine powdery mildew

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Horticulture Research 2023年第5期10卷 139-149页

作者： Surya Sapkota Cheng Zou Craig Ledbetter Anna Underhill Qi Sun David Gadoury Lance Cadle-Davidson School of Integrative Plant Science Cornell AgriTechCornell UniversityGenevaNY14456USA BRC Bioinformatics Facility Institute of BiotechnologyCornell UniversityIthacaNY14853USA United States Department of Agriculture(USDA)-Agricultural Research Service(ARS) Crop DiseasesPests and Genetics Research UnitSan Joaquin Valley Agricultural Sciences CenterParlierCA93648USA USDA-ARS Grape Genetics Research UnitGenevaNY14456USA

Powdery mildew resistance genes restrict infection attempts at different stages of pathogenesis.Here,a strong and rapid powdery mildew resistance phenotype was discovered from vitis amurensis‘PI 588631’that rapidly stopped over 97%of Erysiphe necator conidia,before or immediately after emergence of a secondary hypha from appressoria.This resistance was effective across multiple years of vineyard evaluation on leaves,stems,rachises,and fruit and against a diverse array of E.necator laboratory isolates.Using core genome rhAmpSeq markers,resistance mapped to a single dominant locus(here named REN12)on chromosome 13 near 22.8–27.0 Mb,irrespective of tissue type,explaining up to 86.9%of the phenotypic variation observed on leaves.Shotgun sequencing of recombinant vines using skim-seq technology enabled the locus to be further resolved to a 780 kb region,from 25.15 to 25.93 Mb.RNASeq analysis indicated the allele-specific expression of four resistance genes(NLRs)from the resistant parent.REN12 is one of the strongest powdery mildew resistance loci in grapevine yet documented,and the rhAmpSeq sequences presented here can be directly used for marker-assisted selection or converted to other genotyping platforms.While no virulent isolates were identified among the genetically diverse isolates and wild populations of E.necator tested here,NLR loci like REN12 are often race-specific.Thus,stacking of multiple resistance genes and minimal use of fungicides should enhance the durability of resistance and could enable a 90%reduction in fungicides in low-rainfall climates where few other pathogens attack the foliage or fruit.

关键词： vitis resistance durability

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QTL detection and candidate gene analysis of grape white rot resistance by interspecific grape (vitis vinifera L. × vitis davidii Foex.) crossing

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Horticulture Research 2023年第5期10卷 236-246页

作者： Peng Li Xibei Tan Ruitao Liu Faiz Ur Rahman Jianfu Jiang Lei Sun Xiucai Fan Jihong Liu Chonghuai Liu Ying Zhang National Key Laboratory for Germplasm Innovation and Utilization of Horticultural Crops Zhengzhou Fruit Research InstituteChinese Academy of Agricultural SciencesZhengzhou 450000China Key Laboratory of Horticultural Plant Biology(MOE) College of Horticulture and Forestry SciencesHuazhong Agricultural UniversityWuhan 430000China Zhongyuan Research Center Chinese Academy of Agricultural Sciences

Grape white rot,a devastating disease of grapevines caused by Coniella diplodiella(Speg.)Sacc.,leads to significant yield losses in grape.Breeding grape cultivars resistant to white rot is essential to reduce the regular use of chemical treatments.In recent years,Chinese grape species have gained more attention for grape breeding due to their high tolerance to various biotic and abiotic factors along with changing climatic conditions.In this study,we employed whole-genome resequencing(WGR)to genotype the parents of‘Manicure Finger’(vitis vinifera,female)and‘0940’(vitis davidii,male),along with 101 F1 mapping population individuals,thereby constructing a linkage genetic map.The linkage map contained 9337 single-nucleotide polymorphism(SNP)markers with an average marker distance of 0.3 cM.After 3 years of phenotypic evaluation of the progeny for white rot resistance,we confirmed one stable quantitative trait locus(QTL)for white rot resistance on chromosome 3,explaining up to 17.9%of the phenotypic variation.For this locus,we used RNA-seq to detect candidate gene expression and identified PR1 as a candidate gene involved in white rot resistance.Finally,we demonstrated that recombinant PR1 protein could inhibit the growth of C.diplodiella and that overexpression of PR1 in susceptible V.vinifera increased grape resistance to the pathogen.

关键词： white vitis vinifera

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Divergent molecular and growth responses of young“Cabernet Sauvignon”(vitis vinifera)plants to simple and mixed infections with Grapevine rupestris stem pitting-associated virus

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Horticulture Research 2020年第1期7卷 2706-2719页

作者： M.Tobar N.Fiore A.G.Pérez-Donoso R.León I.M.Rosales M.Gambardella Pontificia Universidad Católica de Chile Facultad de Agronomía e Ingeniería ForestalVicuña Mackena 4860MaculSantiago 7820436Chile Universidad de Chile Facultad de Ciencias AgronómicasAvenida Santa Rosa 11315La PintanaSantiago 8820808Chile

Grapevine rupestris stem pitting associated virus(GRSPaV)is one of the most widely distributed viruses;even so,little is known about its effect on vitis vinifera.To provide new insights,the effects of single and mixed GRSPaV infections on the V.vinifera cultivar“Cabernet Sauvignon”were studied by evaluating growth parameters,such as measurements of the total plant length,the number and distance of internodes and the number of leaves per shoot.In addition,parameters relating to gas exchange,i.e.,the stomatal conductance,net photosynthetic rate,internal CO2 concentration and leaf transpiration,were also assessed.All the measurements were performed in one-and two-yearold plants with a single GRSPaV infection or mixed infections of GRSPaV and Grapevine fanleaf virus(GFLV).The results show that the plant phytosanitary status did not significantly alter the growth and gas exchange parameters in oneyear-old plants.However,in two-year-old plants,single GRSPaV infections increased shoot elongation,which was accompanied by the overexpression of genes associated with the gibberellic acid response pathway.The gas exchange parameters of these plants were negatively affected,despite exhibiting higher LHCII gene expression.Plants with mixed infections did not have modified growth parameters,although they presented a greater reduction in the primary photosynthetic parameters evaluated with no change in LHCII expression.The results presented here confirm the co-evolution hypothesis for V.vinifera and GRSPaV during the early stages of plant development,and they provide new evidence about the effects of GRSPaV and GFLV co-infections on the“Cabernet Sauvignon”cultivar.

关键词： vitis infections vinifera

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CRISPR/Cas9-mediated mutagenesis of VvMLO3 results in enhanced resistance to powdery mildew in grapevine(vitis vinifera)

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Horticulture Research 2020年第1期7卷 1200-1213页

作者： Dong-Yan Wan Ye Guo Yuan Cheng Yang Hu Shunyuan Xiao Yuejin Wang Ying-Qiang Wen State Key Laboratory of Crop Stress Biology for Arid Areas College of HorticultureNorthwest A&F UniversityYangling 712100 ShaanxiChina Key Laboratory of Horticultural Plant Biology and Germplasm Innovation in Northwest China Ministry of AgricultureYangling 712100 ShaanxiChina Institute for Bioscience and Biotechnology Research&Department of Plant Sciences and Landscape Architecture University of Maryland College ParkRockvilleMD 20850USA

Grapevine(vitis vinifera),one of the most economically important fruit crops in the world,suffers significant yield losses from powdery mildew,a major fungal disease caused by Erysiphe necator.In addition to suppressing host immunity,phytopathogens modulate host proteins termed susceptibility(S)factors to promote their proliferation in plants.In this study,CRISPR/Cas9(clustered regularly interspaced short palindromic repeats/CRISPR-associated 9)technology was used to enable the targeted mutagenesis of MLO(mildew resistance Locus O)family genes that are thought to serve as S factors for powdery mildew fungi.Small deletions or insertions were induced in one or both alleles of two grapevine MLO genes,VvMLO3 and VvMLO4,in the transgenic plantlets of the powdery mildew-susceptible cultivar Thompson Seedless.The editing efficiency achieved with different CRISPR/Cas9 constructs varied from 0 to 38.5%.Among the 20 VvMLO3/4-edited lines obtained,one was homozygous for a single mutation,three harbored biallelic mutations,seven were heterozygous for the mutations,and nine were chimeric,as indicated by the presence of more than two mutated alleles in each line.Six of the 20 VvMLO3/4-edited grapevine lines showed normal growth,while the remaining lines exhibited senescence-like chlorosis and necrosis.Importantly,four VvMLO3-edited lines showed enhanced resistance to powdery mildew,which was associated with host cell death,cell wall apposition(CWA)and H2O2 accumulation.Taken together,our results demonstrate that CRISPR/Cas9 genome-editing technology can be successfully used to induce targeted mutations in genes of interest to improve traits of economic importance,such as disease resistance in grapevines.

关键词： resistance vitis insertion

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CRISPR/Cas9-mediated VvPR4b editing decreases downy mildew resistance in grapevine(vitis vinifera L.)

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Horticulture Research 2020年第1期7卷 979-989页

作者： Meng-Yuan Li Yun-Tong Jiao Yu-Ting Wang Na Zhang Bian-Bian Wang Rui-Qi Liu Xiao Yin Yan Xu Guo-Tian Liu State Key Laboratory of Crop Stress Biology in Arid Areas College of HorticultureNorthwest A&F UniversityYanglingShaanxi 712100China Key Laboratory of Horticultural Plant Biology and Germplasm Innovation in Northwest China Ministry of AgricultureNorthwest A&F UniversityYanglingShaanxi 712100China

Downy mildew of grapevine(vitis vinifera L.),caused by the oomycete pathogen Plasmopara viticola,is one of the most serious concerns for grape production worldwide.It has been widely reported that the pathogenesis-related 4(PR4)protein plays important roles in plant resistance to diseases.However,little is known about the role of PR4 in the defense of grapevine against P.viticola.In this study,we engineered loss-of-function mutations in the VvPR4b gene from the cultivar“Thompson Seedless”using the CRISPR/Cas9 system and evaluated the consequences for downy mildew resistance.Sequencing results showed that deletions were the main type of mutation introduced and that no off-target events occurred.Infection assays using leaf discs showed that,compared to wild-type plants,the VvPR4b knockout lines had increased susceptibility to P.viticola.This was accompanied by reduced accumulation of reactive oxygen species around stomata.Measurement of the relative genomic abundance of P.viticola in VvPR4b knockout lines also demonstrated that the mutants had increased susceptibility to the pathogen.Our results confirm that VvPR4b plays an active role in the defense of grapevine against downy mildew.

关键词： CRISPR/Cas9 resistance vitis

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Wild grapevine(vitis vinifera subsp.sylvestris)in the Hyrcanian relict forests of northern Iran:an overview of current taxonomy,ecology and palaeorecords

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Journal of Forestry Research 2018年第6期29卷 1757-1768页

作者： Alireza Naqinezhad Elias Ramezani Morteza Djamali Annik Schnitzler Claire Arnold Department of Biology Faculty of Basic SciencesUniversity of MazandaranBabolsarMazandaranIran Department of Forestry Faculty of Natural ResourcesUrmia UniversityUrmiaIran Institut Mediterraneen de Biodiversite et d’Ecologie(IMBE UMR 7263CNRS-Aix Marseille Universite-IRD-Universite d’Avignon)Europole mediterraneen de l'Arboisbatiment VilleminBP8013545 Aix-En-Provence Cedex 04France Laboratoire Interdisciplinaire des Environnements Continentaux LIEC-UMR 7360 CNRSUniversity of LorraineMetzFrance University of Lausanne Unicentre1015 LausanneSwitzerland

Due to severe anthropogenic impacts on lowland and submontane zones of the Hyrcanian forests of northern Iran, wild grapevine(vitis vinifera subsp. sylvestris), a sporadically distributed woody liana, is currently considered an endangered species. Using data from the literature and 34 studied populations, herbarium assessments and nine palynological sites, we provide an overview of its taxonomy,distribution and ecology in the first part of the investigation.The separation of the two subspecies, namely V. sylvestris subsp. anebophylla and V. sylvestris subsp. trichophylla(sensu Flora Iranica), based on their leaf indumentums, could not be confirmed by our examination of herbarium materials and field observations. Indumentum of the leaves is a result of leaf polymorphism in different vitis specimens and can be strongly influenced by environmental conditions. vitis vinifera subsp. sylvestris grows in a wide range of habitats including wetlands, seasonal stream sides in closed forests,alluvial beds of large rivers, sand dune shrublands and forested wetlands(alder forests). Parrotia persica and Carpinus betulus were the most frequent host species in the studied populations. In the Hyrcanian region, no pollen record of vitis older than the Mid-or Late-Holocene has yet been established. Despite the intrinsic problem in pollen identification by normal(i.e., light) microscopy of wild from cultivated grapevines, the significant values and persistent occurrence of vitis pollen since the Mid-Holocene(before the Bronze Age) in the Hyrcanian pollen records may imply the onset of viticulture in low-to mid-elevation sites in the region. This represents an argument to consider the Hyrcanian region as a possible domestication center for V. vinifera.However, the question of wild versus cultivated origin of grapevines in the Hyrcanian pollen records and the possible date of its domestication and/or cultivation will remain open until further palynological studies are undertaken.

关键词： Domestication Grapevine Hyrcanian forests Pollen vitis

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Effects of exogenous paclobutrazol and sampling time on the efficiency of in vitro embryo rescue in the breeding of new seedless grape varieties

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Journal of Integrative Agriculture 2022年第6期21卷 1633-1644页

作者： XU Teng-fei GUO Yu-rui WANG Wen-yuan YUAN Xiao-jian CHU Yan-nan WANG Xiao-wei HAN Yu-lei WANG Yue-jin SONG Rui FANG Yu-lin WANG Lu-jun XU Yan State Key Laboratory of Crop Stress Biology for Arid Areas College of HorticultureNorthwest A&F UniversityYangling 712100P.R.China College of Plant Science Tibet Agricultural and Animal Husbandry UniversityNyingchi 860000P.R.China Institute for China Wine Industry Technology Yinchuan 750000P.R.China Institute for Weinan Grape Research Weinan 714099P.R.China

Embryo rescue technology plays an important role in seedless grape breeding.However,the efficiency of embryo rescue,including the embryo formation,germination,and seedling rates,is closely related to the parental genotypes,degree of abortion,growth medium,and plant growth regulators.In this study,we investigated the effects of different concentrations of paclobutrazol(PAC),a plant growth regulator,and embryo collection times on the embryo formation,germination,and seedling rates for different hybrid combinations of grape breeding varieties used for their aroma and cold-resistance traits.The results showed that the different PAC concentrations had varying impacts on the development of ovules and embryos from the different grape varieties.The embryo formation rates of the‘Sultanina Rose’×‘Beibinghong’and‘Kunxiang Seedless’×‘Taishan-2’crosses were the highest under the 5.1μmol L^(-1)PAC treatment.The 1.0μmol L^(-1)PAC treatment was optimal for the germination and seedling development of the‘Sultanina Rose’×‘Beibinghong’embryos,whereas the 0.2μmol L^(-1)PAC treatment induced the highest germination rate for the‘Sultanina Rose’×‘Kunxiang Seedless’cross.The optimal sampling times for each cross varied as 39 d after pollination(DAP)for the‘Flame Seedless’×‘Muscat Hamburg’cross,46 DAP for the‘Kunxiang Seedless’×‘Beibinghong’cross,and 41 DAP for the‘Ruby Seedless’×‘Beibinghong’and‘Fantasy Seedless’×‘Shuangyou’crosses.Moreover,the medium modified with 0.5 g L^(-1)of indole-3-butyric acid allowed the malformed seedlings to develop into plantlets and achieve larger progenies.This study provides a useful basis for further studies into grape embryo rescue and could improve breeding efforts for new seedless grape varieties.

关键词： vitis grape embryo rescue paclobutrazol malformed seedlings

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Identification of potential key genes in resveratrol biosynthesis via transcriptional analyses of berry development in grapevine(vitis spp.)genotypes varying in trans-resveratrol content

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Fruit Research 2022年第1期2卷 52-61页

作者： Xiuming Zhang Yifei Wu Yichu Zhang Xiangjing Yin Steve van Nocker Junqiang Guo Zhi Li Min Gao Junyang Song Xiping Wang State Key Laboratory of Crop Stress Biology in Arid Areas College of HorticultureNorthwest A&F UniversityYangling 712100China Key Laboratory of Horticultural Plant Biology and Germplasm Innovation in Northwest China Ministry of AgricultureNorthwest A&F UniversityYangling 712100China Department of Horticulture Michigan State UniversityEast Lansing 48824USA

Resveratrol is an important secondary metabolite not only owing to its function as a phytoalexin,but also its potential benefits to human health.In this study,the content of trans-resveratrol was documented in seven accessions of grapevine,in the seed,pulp and skin of berries,and at three developmental stages.The highest amount(2.99μg g^(−1) FW)was found in the skin of berries at the ripe stage from V.amurensis‘Tonghua-3'.Resveratrol was not detected in several samples,including skin of berries at the green hard or véraison stage from V.davidii‘Tangwei'.We carried out transcriptional profiling of developing‘Tonghua-3'and‘Tangwei'berries to identify gene expression patterns that may be linked with the difference in resveratrol content between these accessions.The expression levels of several differentially expressed genes(DEGs)with presumed function in resveratrol biosynthesis,including STILBENE SYNTHASEs(STSs),CINNAMATE 4-HYDROXYLASEs(C4Hs)and 4-COUMARATE-COA LIGASEs(4CLs),were significantly higher in‘Tonghua-3',than in'Tangwei'during the véraison and ripe stages.Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses suggested that these DEGs were enriched for multiple biological processes at the three stages of fruit development.Additionally,we identified a total of 36 transcription factors,including MYBs,WRKYs,ERFs,bHLHs and bZIPs,that were coexpressed with 17 STSs via a weighted gene co-expression network analysis,suggesting roles as regulators of resveratrol biosynthesis.Overall,these findings provide insight into genotypic differences in resveratrol biosynthesis in grapevine,as well as the molecular genetics of its regulation.

关键词： vitis content potential

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An optimized procedure for plant recovery from somatic embryos significantly facilitates the genetic improvement of vitis

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Horticulture Research 2014年第1期1卷 183-189页

作者： Zhijian T Li Kyung-Hee Kim Sadanand A Dhekney Jonathan R Jasinski Matthew R Creech Dennis J Gray Grape Biotechnology Core Laboratory Mid-Florida Research and Education CenterUniversity of Florida/IFASApopkaFL 32703-8504USA Department of Plant Sciences Sheridan Research and Extension CenterSheridanWY 82801USA

Plant regeneration from grapevine(vitis spp.)via somatic embryogenesis typically is poor.Recovery of plants from vitis rotundifolia Michx.(muscadine grape)is particularly problematic due to extremely low efficiency,including extended culture durations required for embryo–plant conversion.Poor plant recovery is an obstacle to the selection of improved genetically modified lines.Somatic embryos(SEs)of V.rotundifolia cultivar Delicious(Del-HS)and vitis vinifera L cultivar Thompson Seedless(TS)were used to identify culture media and conditions that promoted embryo differentiation and plant conversion;this resulted in a two-step culture system.In comparative culture experiments,C2D medium containing 6%sucrose was the most effective,among four distinct formulae tested,for inducing precocious SE germination and cell differentiation.This medium,further supplemented with 4 mM 6-benzylaminopurine(C2D4B),was subsequently determined to enhance post-germinative growth of SE.MS medium supplemented with 0.5 mM 1-naphthaleneacetic acid(MSN)was then utilized to stimulate root and shoot growth of germinated SE.An average of 35%and 80%‘Del-HS’and‘TS’SE,respectively,developed into plants.All plants developed robust root and shoot systems and exhibited excellent survival following transfer to soil.Over 150 plants of‘Del-HS’were regenerated and established within 2.5 months,which is a dramatic reduction from the 6-to 12-month time period previously required.Similarly,88‘TS’plant lines were obtained within the same time period.Subsequently,seven out of eight vitis cultivars exhibited significantly increased plant conversion percentages,demonstrating broad application of the two-step culture system to produce the large numbers of independent plant lines needed for selection of desired traits.

关键词： cultivar vitis culture

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