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    Predicting community traits along an alpine grassland transect using field imaging spectroscopy
    Feng Zhang, Wenjuan Wu, Lang Li, Xiaodi Liu, Guangsheng Zhou and Zhenzhu Xu
    J Integr Plant Biol 2023, 65 (12): 2604-2618.  
    DOI: 10.1111/jipb.13572
    Abstract (Browse 134)  |   Save
    Assessing plant community traits is important for understanding how terrestrial ecosystems respond and adapt to global climate change. Field hyperspectral remote sensing is effective for quantitatively estimating vegetation properties in most terrestrial ecosystems, although it remains to be tested in areas with dwarf and sparse vegetation, such as the Tibetan Plateau. We measured canopy reflectance in the Tibetan Plateau using a handheld imaging spectrometer and conducted plant community investigations along an alpine grassland transect. We estimated community structural and functional traits, as well as community function based on a field survey and laboratory analysis using 14 spectral vegetation indices (VIs) derived from hyperspectral images. We quantified the contributions of environmental drivers, VIs, and community traits to community function by structural equation modelling (SEM). Univariate linear regression analysis showed that plant community traits are best predicted by the normalized difference vegetation index, enhanced vegetation index, and simple ratio. Structural equation modelling showed that VIs and community traits positively affected community function, whereas environmental drivers and specific leaf area had the opposite effect. Additionally, VIs integrated with environmental drivers were indirectly linked to community function by characterizing the variations in community structural and functional traits. This study demonstrates that community-level spectral reflectance will help scale plant trait information measured at the leaf level to larger-scale ecological processes. Field imaging spectroscopy represents a promising tool to predict the responses of alpine grassland communities to climate change.
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    Efficient CRISPR/Cas9-mediated genome editing in sheepgrass (Leymus chinensis)
    Zhelong Lin, Lei Chen, Shanjie Tang, Mengjie Zhao, Tong Li, Jia You, Changqing You, Boshu Li, Qinghua Zhao, Dongmei Zhang, Jianli Wang, Zhongbao Shen, Xianwei Song, Shuaibin Zhang and Xiaofeng Cao
    J Integr Plant Biol 2023, 65 (11): 2416-2420.  
    DOI: 10.1111/jipb.13567
    Abstract (Browse 234)  |   Save
    The lack of genome editing platforms has hampered efforts to study and improve forage crops that can be grown on lands not suited to other crops. Here, we established efficient Agrobacterium-mediated clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated nuclease 9 (Cas9) genome editing in a perennial, stress-tolerant forage grass, sheepgrass (Leymus chinensis). By screening for active single-guide RNAs (sgRNAs), accessions that regenerate well, suitable Agrobacterium strains, and optimal culture media, and co-expressing the morphogenic factor TaWOX5, we achieved 11% transformation and 5.83% editing efficiency in sheepgrass. Knocking out Teosinte Branched1 (TB1) significantly increased tiller number and biomass. This study opens avenues for studying gene function and breeding in sheepgrass.
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    Cas9-targeted Nanopore sequencing rapidly elucidates the transposition preferences and DNA methylation profiles of mobile elements in plants
    Pavel Merkulov, Sofya Gvaramiya, Maxim Dudnikov, Roman Komakhin, Murad Omarov, Alina Kocheshkova, Zakhar Konstantinov, Alexander Soloviev, Gennady Karlov, Mikhail Divashuk and Ilya Kirov
    J Integr Plant Biol 2023, 65 (10): 2242-2261.  
    DOI: 10.1111/jipb.13555
    Abstract (Browse 512)  |   Save
    Transposable element insertions (TEIs) are an important source of genomic innovation by contributing to plant adaptation, speciation, and the production of new varieties. The often large, complex plant genomes make identifying TEIs from short reads difficult and expensive. Moreover, rare somatic insertions that reflect mobilome dynamics are difficult to track using short reads. To address these challenges, we combined Cas9-targeted Nanopore sequencing (CANS) with the novel pipeline NanoCasTE to trace both genetically inherited and somatic TEIs in plants. We performed CANS of the EVADÉ (EVD) retrotransposon in wild-type Arabidopsis thaliana and rapidly obtained up to 40×sequence coverage. Analysis of hemizygous T-DNA insertion sites and genetically inherited insertions of the EVD transposon in the ddm1 (decrease in DNA methylation 1) genome uncovered the crucial role of DNA methylation in shaping EVD insertion preference. We also investigated somatic transposition events of the ONSEN transposon family, finding that genes that are downregulated during heat stress are preferentially targeted by ONSENs. Finally, we detected hypomethylation of novel somatic insertions for two ONSENs. CANS and NanoCasTE are effective tools for detecting TEIs and exploring mobilome organization in plants in response to stress and in different genetic backgrounds, as well as screening T-DNA insertion mutants and transgenic plants.
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    The transcription factor NAC102 confers cadmium tolerance by regulating WAKL11 expression and cell wall pectin metabolism in Arabidopsis
    Guang Hao Han, Ru Nan Huang, Li Hong Hong, Jia Xi Xu, Yi Guo Hong, Yu Huan Wu and Wei Wei Chen
    J Integr Plant Biol 2023, 65 (10): 2262-2278.  
    DOI: 10.1111/jipb.13557
    Abstract (Browse 250)  |   Save
    Cadmium (Cd) toxicity severely limits plant growth and development. Moreover, Cd accumulation in vegetables, fruits, and food crops poses health risks to animals and humans. Although the root cell wall has been implicated in Cd stress in plants, whether Cd binding by cell wall polysaccharides contributes to tolerance remains controversial, and the mechanism underlying transcriptional regulation of cell wall polysaccharide biosynthesis in response to Cd stress is unknown. Here, we functionally characterized an Arabidopsis thaliana NAC-type transcription factor, NAC102, revealing its role in Cd stress responses. Cd stress rapidly induced accumulation of NAC102.1, the major transcript encoding functional NAC102, especially in the root apex. Compared to wild type (WT) plants, a nac102 mutant exhibited enhanced Cd sensitivity, whereas NAC102.1-overexpressing plants displayed the opposite phenotype. Furthermore, NAC102 localizes to the nucleus, binds directly to the promoter of WALL-ASSOCIATED KINASE-LIKE PROTEIN11 (WAKL11), and induces transcription, thereby facilitating pectin degradation and decreasing Cd binding by pectin. Moreover, WAKL11 overexpression restored Cd tolerance in nac102 mutants to the WT levels, which was correlated with a lower pectin content and lower levels of pectin-bound Cd. Taken together, our work shows that the NAC102-WAKL11 module regulates cell wall pectin metabolism and Cd binding, thus conferring Cd tolerance in Arabidopsis.
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    A straight-forward seed production technology system for foxtail millet (Setaria italica)
    Wei Zhang, Xiantao Qi, Hui Zhi, Yushuang Ren, Linlin Zhang, Yuanzhu Gao, Yi Sui, Haoshan Zhang, Sha Tang, Guanqing Jia, Chuanxiao Xie, Chuanyin Wu and Xianmin Diao
    J Integr Plant Biol 2023, 65 (9): 2023-2035.  
    DOI: 10.1111/jipb.13503
    Abstract (Browse 241)  |   Save
    For autogamous crops, a precondition for using heterosis is to produce sufficient pure male-sterile female parents that can be used to produce hybrid seeds. To date, cytoplasmic male sterility (CMS) and environment-sensitive genic male sterility (EGMS) have been used commercially to exploit heterosis for autogamous species. However, neither CMS nor EGMS has been established for foxtail millet (Setaria italica). Here, we report on the establishment and application of a seed production technology (SPT) system for this crop. First, we established a DsRed-based SPT system, but found that it was unsuitable because it required the use of a fluorescent device for seed sorting. Instead, we constructed an SPT system with de novo betalain biosynthesis as the selection marker. This allowed us to distinguish transgenic seeds with the naked eye, thereby facilitating the identification of SPT maintainer line seeds. In this system, a seed sorter was not required to obtain sufficient seeds. The key point of the strategy is that the seed pool of the SPT maintainer line is propagated by artificial identification and harvesting of male-fertile individuals in the field, and the male-sterile line seed pool for hybrid production is produced and propagated by free pollination of male-sterile plants with the SPT maintainer line. In a field experiment, we obtained 423.96 kg male-sterile line seeds per acre, which is sufficient to plant 700.18 acres of farmland for hybrid seed production or male-sterile line reproduction. Our study therefore describes a powerful tool for hybrid seed production in foxtail millet, and demonstrates how the SPT system can be used for a small-grained crop with high reproduction efficiency.
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    Cited: Web of Science(2)
      
    Integrating multiplicate data: A new trend for taxonomic study
    De‐Yuan Hong
    J Integr Plant Biol 2023, 65 (9): 2021-2022.  
    doi: 10.1111/jipb.13548
    Abstract (Browse 104)  |   Save
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    Development of an efficient expression system with large cargo capacity for interrogation of gene function in bamboo based on bamboo mosaic virus
    Yandong Jin, Baijie Wang, Mingchuan Bao, Yujie Li, Shengwu Xiao, Yuhua Wang, Jun Zhang, Liangzhen Zhao, Hangxiao Zhang, Yau-Heiu Hsu, Mingjie Li and Lianfeng Gu
    J Integr Plant Biol 2023, 65 (6): 1369-1382.  
    DOI: 10.1111/jipb.13468
    Abstract (Browse 225)  |   Save
    Bamboo is one of the fastest growing plants among monocotyledonous species and is grown extensively in subtropical regions. Although bamboo has high economic value and produces much biomass quickly, gene functional research is hindered by the low efficiency of genetic transformation in this species. We therefore explored the potential of a bamboo mosaic virus (BaMV)- mediated expression system to investigate genotype-phenotype associations. We determined that the sites between the triple gene block proteins (TGBps) and the coat protein (CP) of BaMV are the most efficient insertion sites for the expression of exogenous genes in both monopodial and sympodial bamboo species. Moreover, we validated this system by individually overexpressing the two endogenous genes ACE1 and DEC1, which resulted in the promotion and suppression of internode elongation, respectively. In particular, this system was able to drive the expression of three 2A-linked betalain biosynthesis genes (more than 4 kb in length) to produce betalain, indicating that it has high cargo capacity and may provide the prerequisite basis for the development of a DNA-free bamboo genome editing platform in the future. Since BaMV can infect multiple bamboo species, we anticipate that the system described in this study will greatly contribute to gene function research and further promote the molecular breeding of bamboo.
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    A high-resolution transcriptomic atlas depicting nitrogen fixation and nodule development in soybean
    Baocheng Sun, Yu Wang, Qun Yang, Han Gao, Haiyu Niu, Yansong Li, Qun Ma, Qing Huan, Wenfeng Qian and Bo Ren
    J Integr Plant Biol 2023, 65 (6): 1536-1552.  
    doi: 10.1111/jipb.13495
    Abstract (Browse 256)  |   Save
    Although root nodules are essential for biological nitrogen fixation in legumes, the cell types and molecular regulatory mechanisms contributing to nodule development and nitrogen fixation in determinate nodule legumes, such as soybean (Glycine max), remain incompletely understood. Here, we generated a single-nucleus resolution transcriptomic atlas of soybean roots and nodules at 14 days post inoculation (dpi) and annotated 17 major cell types, including six that are specific to nodules. We identified the specific cell types responsible for each step in the ureides synthesis pathway, which enables spatial compartmentalization of biochemical reactions during soybean nitrogen fixation. By utilizing RNA velocity analysis, we reconstructed the differentiation dynamics of soybean nodules, which differs from those of indeterminate nodules in Medicago truncatula. Moreover, we identified several putative regulators of soybean nodulation and two of these genes, GmbHLH93 and GmSCL1, were as-yet uncharacterized in soybean. Overexpression of each gene in soybean hairy root systems validated their respective roles in nodulation. Notably, enrichment for cytokinin-related genes in soybean nodules led to identification of the cytokinin receptor, GmCRE1, as a prominent component of the nodulation pathway. GmCRE1 knockout in soybean resulted in a striking nodule phenotype with decreased nitrogen fixation zone and depletion of leghemoglobins, accompanied by downregulation of nodule-specific gene expression, as well as almost complete abrogation of biological nitrogen fixation. In summary, this study provides a comprehensive perspective of the cellular landscape during soybean nodulation, shedding light on the underlying metabolic and developmental mechanisms of soybean nodule formation.
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    Single‐cell RNA‐seq of Lotus japonicus provide insights into identification and function of root cell types of legume
    Zhanmin Sun, Sanjie Jiang, Dan Wang, Linxia Li, Boxin Liu, Qifan Ran, Lizhen Hu, Junbo Xiong, Yixiong Tang, Xiaofeng Gu, Yanmin Wu and Zhe Liang
    J Integr Plant Biol 2023, 65 (5): 1147-1152.  
    DOI: 10.1111/jipb.13435
    Abstract (Browse 230)  |   Save
    The roots of legume plant play a crucial role in nitrogen fixation. However, the transcriptomes of different cell types of legume root and their functions remain largely unknown. Here, we performed single‐cell RNA sequencing and profiled more than 22,000 single cells from root tips of Lotus japonicus, a model species of legume. We identified seven clusters corresponding to seven major cell types, which were validated by in situ hybridization. Further analysis revealed regulatory programs including phytohormone and nodulation associated with specific cell types, and revealed conserved and diverged features for the cell types. Our results represent the first single‐cell resolution transcriptome for legume root tips and a valuable resource for studying the developmental and physiological functions of various cell types in legumes.
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    Efficient genotype-independent cotton genetic transformation and genome editing
    Xiaoyang Ge, Jieting Xu, Zhaoen Yang, Xiaofeng Yang, Ye Wang, Yanli Chen, Peng Wang and Fuguang Li
    J Integr Plant Biol 2023, 65 (4): 907-917.  
    doi: 10.1111/jipb.13427
    Abstract (Browse 443)  |   Save
    Cotton (Gossypium spp.) is one of the most important fiber crops worldwide. In the last two decades, transgenesis and genome editing have played important roles in cotton improvement. However, genotype dependence is one of the key bottlenecks in generating transgenic and gene‐edited cotton plants through either particle bombardment or Agrobacterium‐mediated transformation. Here, we developed a shoot apical meristem (SAM) cell‐ mediated transformation system (SAMT) that allowed the transformation of recalcitrant cotton genotypes including widely grown upland cotton (Gossypium hirsutum), Sea island cotton (Gossypium barbadense), and Asiatic cotton (Gossypium arboreum). Through SAMT, we successfully introduced two foreign genes, GFP and RUBY, into SAM cells of some recalcitrant cotton genotypes. Within 2–3 months, transgenic adventitious shoots generated from the axillary meristem zone could be recovered and grown into whole cotton plants. The GFP fluorescent signal and betalain accumulation could be observed in various tissues in GFP‐ and RUBY‐positive plants, as well as in their progenies, indicating that the transgenes were stably integrated into the genome and transmitted to the next generation. Furthermore, using SAMT, we successfully generated edited cotton plants with inheritable targeted mutagenesis in the GhPGF and GhRCD1 genes through CRISPR/Cas9‐mediated genome editing. In summary, the established SAMT transformation system here in this study bypasses the embryogenesis process during tissue culture in a conventional transformation procedure and significantly accelerates the generation of transgenic and gene‐edited plants for genetic improvement of recalcitrant cotton varieties.
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    Cited: Web of Science(12)
      
    Optimized prime editing efficiently generates heritable mutations in maize
    Dexin Qiao, Junya Wang, Min‐Hui Lu, Cuiping Xin, Yiping Chai, Yuanyuan Jiang, Wei Sun, Zhenghong Cao, Siyi Guo, Xue‐Chen Wang and Qi‐Jun Chen
    J Integr Plant Biol 2023, 65 (4): 900-906.  
    doi: 10.1111/jipb.13428
    Abstract (Browse 258)  |   Save
    Low efficiency is the main obstacle to using prime editing in maize (Zea mays). Recently, prime‐editing efficiency was greatly improved in mammalian cells and rice (Oryza sativa) plants by engineering prime‐ editing guide RNAs (pegRNAs), optimizing the prime editor (PE) protein, and manipulating cellular determinants of prime editing. In this study, we tested PEs optimized via these three strategies in maize. We demonstrated that the ePE5max system, composed of PEmax, epegRNAs (pegRNA‐ evopreQ. 1), nicking single guide RNAs (sgRNAs), and MLH1dn, efficiently generated heritable mutations that conferred resistance to herbicides that inhibit 5‐enolpyruvylshikimate‐3‐phosphate synthase (EPSPS), acetolactate synthase (ALS), or acetyl CoA carboxylase (ACCase) activity. Collectively, we demonstrate that the ePE5max system has sufficient efficiency to generate heritable (homozygous or heterozygous) mutations in maize target genes and that the main obstacle to using PEs in maize has thus been removed.
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    Cited: Web of Science(6)
      
    A prolific and robust whole-genome genotyping method using PCR amplification via primer-template mismatched annealing
    Sheng Zhao, Cuicui Zhang, Liqun Wang, Minxuan Luo, Peng Zhang, Yue Wang, Waqar Afzal Malik, Yue Wang, Peng Chen, Xianjin Qiu, Chongrong Wang, Hong Lu, Yong Xiang, Yuwen Liu, Jue Ruan, Qian Qian, Haijian Zhi and Yuxiao Chang
    J Integr Plant Biol 2023, 65 (3): 633-645.  
    doi: 10.1111/jipb.13395
    Abstract (Browse 382)  |   Save
    Whole-genome genotyping methods are important for breeding. However, it has been challenging to develop a robust method for simultaneous foreground and background genotyping that can easily be adapted to different genes and species. In our study, we accidently discovered that in adapter ligation-mediated PCR, the amplification by primer-template mismatched annealing (PTMA) along the genome could generate thousands of stable PCR products. Based on this observation, we consequently developed a novel method for simultaneous foreground and background integrated genotyping by sequencing (FBI-seq) using one specific primer, in which foreground genotyping is performed by primer-template perfect annealing (PTPA), while background genotyping employs PTMA. Unlike DNA arrays, multiple PCR, or genome target enrichments, FBI-seq requires little preliminary work for primer design and synthesis, and it is easily adaptable to different foreground genes and species. FBI-seq therefore provides a prolific, robust, and accurate method for simultaneous foreground and background genotyping to facilitate breeding in the post-genomics era.
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    Cited: Web of Science(1)
      
    Time series canopy phenotyping enables the identification of genetic variants controlling dynamic phenotypes in soybean
    Delin Li, Dong Bai, Yu Tian, Ying-Hui Li, Chaosen Zhao, Qi Wang, Shiyu Guo, Yongzhe Gu, Xiaoyan Luan, Ruizhen Wang, Jinliang Yang, Malcolm J. Hawkesford, James C. Schnable, Xiuliang Jin, Li-Juan Qiu
    J Integr Plant Biol 2023, 65 (1): 117-132.  
    doi: 10.1111/jipb.13380
    Abstract (Browse 283)  |   Save
    Advances in plant phenotyping technologies are dramatically reducing the marginal costs of collecting multiple phenotypic measurements across several time points. Yet, most current approaches and best statistical practices implemented to link genetic and phenotypic variation in plants have been developed in an era of single-time-point data. Here, we used time-series phenotypic data collected with an unmanned aircraft system for a large panel of soybean (Glycine max (L.) Merr.) varieties to identify previously uncharacterized loci. Specifically, we focused on the dissection of canopy coverage (CC) variation from this rich data set. We also inferred the speed of canopy closure, an additional dimension of CC, from the time-series data, as it may represent an important trait for weed control. Genome-wide association studies (GWASs) identified 35 loci exhibiting dynamic associations with CC across developmental stages. The time-series data enabled the identification of 10 known flowering time and plant height quantitative trait loci (QTLs) detected in previous studies of adult plants and the identification of novel QTLs influencing CC. These novel QTLs were disproportionately likely to act earlier in development, which may explain why they were missed in previous single-time-point studies. Moreover, this time-series data set contributed to the high accuracy of the GWASs, which we evaluated by permutation tests, as evidenced by the repeated identification of loci across multiple time points. Two novel loci showed evidence of adaptive selection during domestication, with different genotypes/haplotypes favored in different geographic regions. In summary, the time-series data, with soybean CC as an example, improved the accuracy and statistical power to dissect the genetic basis of traits and offered a promising opportunity for crop breeding with quantitative growth curves.
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    Cited: Web of Science(7)
      
    A cost-effective tsCUT&Tag method for profiling transcription factor binding landscape
    Leiming Wu, Zi Luo, Yanni Shi, Yizhe Jiang, Ruonan Li, Xinxin Miao, Fang Yang, Qing Li, Han Zhao, Jiquan Xue, Shutu Xu, Tifu Zhang and Lin Li
    J Integr Plant Biol 2022, 64 (11): 2033-2038.  
    doi: 10.1111/jipb.13354
    Abstract (Browse 362)  |   Save

    Knowledge of the transcription factor binding landscape (TFBL) is necessary to analyze gene regulatory networks for important agronomic traits. However, a low-cost and high-throughput in vivo chromatin profiling method is still lacking in plants. Here, we developed a transient and simplified cleavage under targets and tagmentation (tsCUT&Tag) that combines transient expression of transcription factor proteins in protoplasts with a simplified CUT&Tag without nucleus extraction. Our tsCUT&Tag method provided higher data quality and signal resolution with lower sequencing depth compared with traditional ChIP-seq. Furthermore, we developed a strategy combining tsCUT&Tag with machine learning, which has great potential for profiling the TFBL across plant development.

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    Cited: Web of Science(8)
      
    Rapid improvement of rice eating and cooking quality through gene editing toward glutelin as target
    Yihao Yang, Ziyan Shen, Youguang Li, Chenda Xu, Han Xia, Hao Zhuang, Shengyuan Sun, Min Guo and Changjie Yan
    J Integr Plant Biol 2022, 64 (10): 1860-1865.  
    doi: 10.1111/jipb.13334
    Abstract (Browse 355)  |   Save
    Rice eating and cooking quality (ECQ) is a major concern of breeders and consumers, determining market competitiveness worldwide. Rice grain protein content (GPC) is negatively related to ECQ, making it possible to improve ECQ by manipulating GPC. However, GPC is genetically complex and sensitive to environmental conditions; therefore, little progress has been made in traditional breeding for ECQ. Here, we report that CRISPR/Cas9-mediated knockout of genes encoding the grain storage protein glutelin rapidly produced lines with downregulated GPC and improved ECQ. Our finding provides a new strategy for improving rice ECQ.
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    Cited: Web of Science(5)
      
    Cas12a-based on-site, rapid detection of genetically modified crops
    Zhiqiang Duan, Xiaoliang Yang, Xingkun Ji, Ying Chen, Xiaomu Niu, Anping Guo, Jian‐Kang Zhu, Feng Li, Zhaobo Lang and Hui Zhao
    J Integr Plant Biol 2022, 64 (10): 1856-1859.  
    doi: 10.1111/jipb.13342
    Abstract (Browse 392)  |   Save
    A CRISPR/LbCas12a-based nucleic acid detection method that uses crude leaf extracts as samples and is rapid (≤40 min for a full run) and highly sensitive (0.01%) can be used to monitor genetically modified organisms in the field.
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    Cited: Web of Science(2)
      
    Establishment of a dmp based maternal haploid induction system for polyploid Brassica napus and Nicotiana tabacum
    Yu Zhong, Yuwen Wang, Baojian Chen, Jinchu Liu, Dong Wang, Mengran Li, Xiaolong Qi, Chenxu Liu, Kim Boutilier and Shaojiang Chen
    J Integr Plant Biol 2022, 64 (6): 1281-1294.  
    doi: 10.1111/jipb.13244
    Abstract (Browse 348)  |   Save

    Doubled haploid (DH) technology is used to obtain homozygous lines in a single generation, a technique that significantly accelerates the crop breeding trajectory. Traditionally, in vitro culture is used to generate DHs, but this technique is limited by species and genotype recalcitrance. In vivo haploid induction (HI) through seed is widely and efficiently used in maize and was recently extended to several other crops. Here we show that in vivo HI can be triggered by mutation of DMP maternal haploid inducer genes in allopolyploid (allotetraploid) Brassica napus and Nicotiana tabacum. We developed a pipeline for selection of DMP orthologs for clustered regularly interspaced palindromic repeats mutagenesis and demonstrated average amphihaploid induction rates of 2.4% and 1.2% in multiple B. napus and N. tabacum genotypes, respectively. These results further confirmed the HI ability of DMP gene in polyploid dicot crops. The DMP-HI system offers a novel DH technology to facilitate breeding in these crops. The success of this approach and the conservation of DMP genes in dicots suggest the broad applicability of this technique in other dicot crops.

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    Cited: Web of Science(18)
      
    Testing the polar auxin transport model with a selective plasma membrane H+-ATPase inhibitor
    Yongqing Yang, Xiaohui Liu, Wei Guo, Wei Liu, Wei Shao, Jun Zhao, Junhong Li, Qing Dong, Liang Ma, Qun He, Yingzhang Li, Jianyong Han and Xiaoguang Lei
    J Integr Plant Biol 2022, 64 (6): 1229-1245.  
    DOI: 10.1111/jipb.13256
    Abstract (Browse 251)  |   Save

    Auxin is unique among plant hormones in that its function requires polarized transport across plant cells. A chemiosmotic model was proposed to explain how polar auxin transport is derived by the H+ gradient across the plasma membrane (PM) established by PM H+-adenosine triphosphatases (ATPases). However, a classical genetic approach by mutations in PM H+-ATPase members did not result in the ablation of polar auxin distribution, possibly due to functional redundancy in this gene family. To confirm the crucial role of PM H+-ATPases in the polar auxin transport model, we employed a chemical genetic approach. Through a chemical screen, we identified protonstatin-1 (PS-1), a selective small-molecule inhibitor of PM H+-ATPase activity that inhibits auxin transport. Assays with transgenic plants and yeast strains showed that the activity of PM H+-ATPases affects auxin uptake as well as acropetal and basipetal polar auxin transport. We propose that PS-1 can be used as a tool to interrogate the function of PM H+-ATPases. Our results support the chemiosmotic model in which PM H+-ATPase itself plays a fundamental role in polar auxin transport.

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    Cited: Web of Science(5)
      
    Efficient and genotype independent maize transformation using pollen transfected by DNA-coated magnetic nanoparticles
    Zuo‐Ping Wang, Zhong‐Bao Zhang, Deng‐Yu Zheng, Tong‐Tong Zhang, Xiang‐Long Li, Chun Zhang, Rong Yu, Jian‐Hua Wei and Zhong‐Yi Wu
    J Integr Plant Biol 2022, 64 (6): 1145-1156.  
    doi: 10.1111/jipb.13263
    Abstract (Browse 494)  |   Save

    Current gene delivery methods for maize are limited to specific genotypes and depend on time-consuming and labor-intensive tissue culture techniques. Here, we report a new method to transfect maize that is culture-free and genotype independent. To enhance efficiency of DNA entry and maintain high pollen viability of 32%-55%, transfection was performed at cool temperature using pollen pretreated to open the germination aperture (40%–55%). Magnetic nanoparticles (MNPs) coated with DNA encoding either red fluorescent protein (RFP), β-glucuronidase gene (GUS), enhanced green fluorescent protein (EGFP) or bialaphos resistance (bar) was delivered into pollen grains, and female florets of maize inbred lines were pollinated. Red fluorescence was detected in 22% transfected pollen grains, and GUS stained 55% embryos at 18 d after pollination. Green fluorescence was detected in both silk filaments and immature kernels. The T1 generation of six inbred lines showed considerable EGFP or GUS transcripts (29%–74%) quantitated by polymerase chain reaction, and 5%–16% of the T1 seedlings showed immunologically active EGFP or GUS protein. Moreover, 1.41% of the bar transfected T1 plants were glufosinate resistant, and heritable bar gene was integrated into the maize genome effectively as verified by DNA hybridization. These results demonstrate that exogenous DNA could be delivered efficiently into elite maize inbred lines recalcitrant to tissue culture-mediated transformation and expressed normally through our genotype-independent pollen transfection system.

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    Cited: Web of Science(20)
      
    Creation of fragrant sorghum by CRISPR/Cas9
    Dan Zhang, Sanyuan Tang, Peng Xie, Dekai Yang, Yaorong Wu, Shujing Cheng, Kai Du, Peiyong Xin, Jinfang Chu, Feifei Yu and Qi Xie
    J Integr Plant Biol 2022, 64 (5): 961-964.  
    doi: 10.1111/jipb.13232
    Abstract (Browse 426)  |   Save

    Sorghum, the fifth largest cereal crop, has high value as a staple food and raw material for liquor and vinegar brewing. Due to its high biomass and quality, it is also used as the second most planted silage resource. No fragrant sorghums are currently on the market. Through CRISPR/Cas9-mediated knockout of SbBADH2, we obtained sorghum lines with extraordinary aromatic smell in both seeds and leaves. Animal feeding experiments showed that fragrant sorghum leaves were attractable. We believe this advantage will produce great value in the sorghum market for both grain and whole biomass forage.

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    Cited: Web of Science(13)
      
    SoySNP618K array: A high-resolution single nucleotide polymorphism platform as a valuable genomic resource for soybean genetics and breeding
    Yan‐Fei Li, Ying‐Hui Li, Shan‐Shan Su, Jochen C. Reif, Zhao‐Ming Qi, Xiao‐Bo Wang, Xing Wang, Yu Tian, De‐Lin Li, Ru‐Jian Sun, Zhang‐Xiong Liu, Ze‐Jun Xu, Guang‐Hui Fu, Ya‐Liang Ji, Qing‐Shan Chen, Ji‐Qiang Liu and Li‐Juan Qiu
    J Integr Plant Biol 2022, 64 (3): 632-648.  
    doi: 10.1111/jipb.13202
    Abstract (Browse 240)  |   Save
    Innovations in genomics have enabled the development of low-cost, high-resolution, single nucleotide polymorphism (SNP) genotyping arrays that accelerate breeding progress and support basic research in crop science. Here, we developed and validated the SoySNP618K array (618,888 SNPs) for the important crop soybean. The SNPs were selected from whole-genome resequencing data containing 2,214 diverse soybean accessions; 29.34% of the SNPs mapped to genic regions representing 86.85% of the 56,044 annotated high-confidence genes. Identity-by-state analyses of 318 soybeans revealed 17 redundant accessions, highlighting the potential of the SoySNP618K array in supporting gene bank management. The patterns of population stratification and genomic regions enriched through domestication were highly consistent with previous findings based on resequencing data, suggesting that the ascertainment bias in the SoySNP618K array was largely compensated for. Genome-wide association mapping in combination with reported quantitative trait loci enabled fine-mapping of genes known to influence flowering time, E2 and GmPRR3b, and of a new candidate gene, GmVIP5. Moreover, genomic prediction of flowering and maturity time in 502 recombinant inbred lines was highly accurate (>0.65). Thus, the SoySNP618K array is a valuable genomic tool that can be used to address many questions in applied breeding, germplasm management, and basic crop research.
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    Cited: Web of Science(5)
      
    RNA silencing: From discovery and elucidation to application and perspectives
    Jian‐Hua Zhao and Hui‐Shan Guo
    J Integr Plant Biol 2022, 64 (2): 476-498.  
    doi: 10.1111/jipb.13213
    Abstract (Browse 258)  |   Save
    RNA silencing (or RNA interference, RNAi) is a conserved mechanism for regulating gene expression in eukaryotes. The discovery of natural trans-kingdom RNAi indicated that small RNAs act as signaling molecules and enable communication between organisms in different kingdoms. The phenomenon and potential mechanisms of trans-kingdom RNAi are among the most exciting research topics. To better understand trans-kingdom RNAi, we review the history of the discovery and elucidation of RNAi mechanisms. Based on canonical RNAi mechanisms, we summarize the major points of divergence around RNAi pathways in the main eukaryotes’ kingdoms, including plants, animals, and fungi. We review the representative incidents associated with the mechanisms and applications of trans-kingdom RNAi in crop protection, and discuss the critical factors that should be considered to develop successful trans-kingdom RNAi-based crop protection strategies.
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    Cited: Web of Science(17)
      
    Single-cell RNA sequencing reveals a high-resolution cell atlas of xylem in Populus
    Hui Li, Xinren Dai, Xiong Huang, Mengxuan Xu, Qiao Wang, Xiaojing Yan, Ronald R. Sederoff and Quanzi Li
    J Integr Plant Biol 2021, 63 (11): 1906-1921.  
    DOI: 10.1111/jipb.13159
    Abstract (Browse 1121)  |   Save
    High-throughput single-cell RNA sequencing (scRNA-seq) has advantages over traditional RNA-seq to explore spatiotemporal information on gene dynamic expressions in heterogenous tissues. We performed Drop-seq, a method for the dropwise sequestration of single cells for sequencing, on protoplasts from the differentiating xylem of Populus alba×Populus glandulosa. The scRNA-seq profiled 9,798 cells, which were grouped into 12 clusters. Through characterization of differentially expressed genes in each cluster and RNA in situ hybridizations, we identified vessel cells, fiber cells, ray parenchyma cells and xylem precursor cells. Diffusion pseudotime analyses revealed the differentiating trajectory of vessels, fiber cells and ray parenchyma cells and indicated a different differentiation process between vessels and fiber cells, and a similar differentiation process between fiber cells and ray parenchyma cells. We identified marker genes for each cell type (cluster) and key candidate regulators during developmental stages of xylem cell differentiation. Our study generates a high-resolution expression atlas of wood formation at the single cell level and provides valuable information on wood formation.
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    Establishment of an efficient seed fluorescence reporter‐assisted CRISPR/Cas9 gene editing in maize
    Yuanyuan Yan, Jinjie Zhu, Xiantao Qi, Beijiu Cheng, Changlin Liu and Chuanxiao Xie
    J Integr Plant Biol 2021, 63 (9): 1671-1680.  
    doi: 10.1111/jipb.13086
    Abstract (Browse 439)  |   Save
    Genome editing by clustered regularly interspaced short palindromic sequences (CRISPR)/CRISPR‐associated protein 9 (Cas9) has revolutionized functional gene analysis and genetic improvement. While reporter‐assisted CRISPR/Cas systems can greatly facilitate the selection of genome‐edited plants produced via stable transformation, this approach has not been well established in seed crops. Here, we established the seed fluorescence reporter (SFR)‐assisted CRISPR/Cas9 systems in maize (Zea mays L.), using the red fluorescent DsRED protein expressed in the endosperm (En‐SFR/Cas9), embryos (Em‐SFR/Cas9), or both tissues (Em/En‐SFR/Cas9). All three SFRs showed distinct fluorescent patterns in the seed endosperm and embryo that allowed the selection of seeds carrying the transgene of having segregated the transgene out. We describe several case studies of the implementation of En‐SFR/Cas9, Em‐SFR/Cas9, and Em/En‐ SFR/Cas9 to identify plants not harboring the genome‐editing cassette but carrying the desired mutations at target genes in single genes or in small‐scale mutant libraries, and report on the successful generation of single‐target mutants and/or mutant libraries with En‐SFR/Cas9, Em‐SFR/Cas9, and Em/En‐SFR/Cas9. SFR‐assisted genome editing may have particular value for application scenarios with a low transformation frequency and may be extended to other important monocot seed crops.
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    Efficient generation of homozygous substitutions in rice in one generation utilizing an rABE8e base editor
    Chuang Wei, Chong Wang, Meng Jia, Hong‐Xuan Guo, Peng‐Yu Luo, Mu‐Gui Wang, Jian‐Kang Zhu and Hui Zhang
    J Integr Plant Biol 2021, 63 (9): 1595-1599.  
    doi: 10.1111/jipb.13089
    Abstract (Browse 475)  |   Save
    A new deaminase, TadA8e, was recently evolved in the laboratory. TadA8e catalyzes DNA deamination over 1,000 times faster than ABE7.10. We developed a high-efficiency adenine base editor, rABE8e (rice ABE8e), combining monomeric TadA8e, bis-bpNLS and codon optimization. rABE8e had substantially increased editing efficiencies at NG-protospacer adjacent motif (PAM) and NGG-PAM target sequences compared with ABEmax. For most targets, rABE8e exhibited nearly 100% editing efficiency and high homozygous substitution rates in the specific editing window, especially at Positions A5 and A6. The ability to rapidly generate plant materials with homozygous base substitutions will benefit gene function research and precision molecular breeding.
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    Development of an efficient plant dual cytosine and adenine editor
    Rongfang Xu, Fanna Kong, Ruiying Qin, Juan Li, Xiaoshuang Liu and Pengcheng Wei
    J Integr Plant Biol 2021, 63 (9): 1600-1605.  
    doi: 10.1111/jipb.13146
    Abstract (Browse 380)  |   Save
    An enhanced CDA-like (eCDAL) was established from Japanese lamprey CDA1-like 4 to achieve a high editing frequency in a broad region as a C-terminal cytosine base editors (CT-CBE). Then, a novel plant dual-base editor version 1(pDuBE1) was developed by integrating TadA-8e into eCDAL. The editing efficiency of pDuBE1 could reach to 87.6%, with frequencies of concurrent A-to-G and C-to-T conversions as high as 49.7% in stably transformed plant cells. Our results showed that pDuBE1 could mediate robust dual editing in plant genome, providing a powerful manipulation tool for precise crop breeding and screening platforms for in planta direct evolution.
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    Simultaneous gene editing of three homoeoalleles in self-incompatible allohexaploid grasses
    Liang Zhang, Tao Wang, Guangyang Wang, Aoyue Bi, Misganaw Wassie, Yan Xie, Liwen Cao, Huawei Xu, Jinmin Fu, Liang Chen, Yang Zhao and Tao Hu
    J Integr Plant Biol 2021, 63 (8): 1410-1415.  
    doi: 10.1111/jipb.13101
    Abstract (Browse 295)  |   Save
    Clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system has been widely used for precise gene editing in plants. However, simultaneous gene editing of multiple homoeoalleles remains challenging, especially in self-incompatible polyploid plants. Here, we simultaneously introduced targeted mutations in all three homoeoalleles of two genes in the self-incompatible allohexaploid tall fescue, using both CRISPR/Cas9 and LbCas12a (LbCpf1) systems. Loss-of-function mutants of FaPDS exhibited albino leaves, while knockout of FaHSP17.9 resulted in impaired heat resistance in T0 generation of tall fescue. Moreover, these mutations were inheritable. Our findings demonstrate the feasibility of generating loss-of-function mutants in T0 generation polyploid perennial grasses using CRISPR/Cas systems.
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    Natural variation in linalool metabolites: One genetic locus, many functions?
    Jun He, Rayko Halitschke, Ian T. Baldwin and Meredith C. Schuman
    J Integr Plant Biol 2021, 63 (8): 1416-1421.  
    doi: 10.1111/jipb.13104
    Abstract (Browse 214)  |   Save
    The ubiquitous volatile linalool is metabolized in plants to nonvolatile derivatives. We studied Nicotiana attenuata plants which naturally vary in (S)-(+)-linalool contents, and lines engineered to produce either (R)-(-)- or (S)-(+)-linalool. Only (S)-(+)-linalool production was associated with slower growth of a generalist herbivore, and a large fraction was present as nonvolatile derivatives. We found that variation in volatile linalool and its nonvolatile glycosides mapped to the same genetic locus which harbored the biosynthetic gene, NaLIS, but that free linalool varied more in environmental responses. This study reveals how (S)-(+)-linalool and conjugates differ in their regulation and possible functions in resistance.
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    LjaFGD: Lonicera japonica functional genomics database
    Qiaoqiao Xiao, Zhongqiu Li, Mengmeng Qu, Wenying Xu, Zhen Su and Jiaotong Yang
    J Integr Plant Biol 2021, 63 (8): 1422-1436.  
    DOI: 10.1111/jipb.13112
    Abstract (Browse 259)  |   Save
    Lonicera japonica Thunb., a traditional Chinese herb, has been used for treating human diseases for thousands of years. Recently, the genome of L. japonica has been decoded, providing valuable information for research into gene function. However, no comprehensive database for gene functional analysis and mining is available for L. japonica. We therefore constructed LjaFGD (www.gzybioinformatics.cn/LjaFGD and bioinformatics.cau.edu.cn/LjaFGD), a database for analyzing and comparing gene function in L. japonica. We constructed a gene co-expression network based on 77 RNA-seq samples, and then annotated genes of L. japonica by alignment against protein sequences from public databases. We also introduced several tools for gene functional analysis, including Blast, motif analysis, gene set enrichment analysis, heatmap analysis, and JBrowse. Our co-expression network revealed that MYB and WRKY transcription factor family genes were co-expressed with genes encoding key enzymes in the biosynthesis of chlorogenic acid and luteolin in L. japonica. We used flavonol synthase 1 (LjFLS1) as an example to show the reliability and applicability of our database. LjaFGD and its various associated tools will provide researchers with an accessible platform for retrieving functional information on L. japonica genes to further biological discovery.
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    Plant-based, adjuvant-free, potent multivalent vaccines for avian influenza virus via Lactococcus surface display
    Shi‐Jian Song, Gyeong‐Im Shin, Jinyong Noh, Jiho Lee, Deok‐Hwan Kim, Gyeongryul Ryu, Gyeongik Ahn, Hyungmin Jeon, Hai‐Ping Diao, Youngmin Park, Min Gab Kim, Woe‐Yeon Kim, Young‐Jin Kim, Eun‐Ju Sohn, Chang Seon Song and Inhwan Hwang
    J Integr Plant Biol 2021, 63 (8): 1505-1520.  
    DOI: 10.1111/jipb.13141
    Abstract (Browse 288)  |   Save
    Influenza epidemics frequently and unpredictably break out all over the world, and seriously affect the breeding industry and human activity. Inactivated and live attenuated viruses have been used as protective vaccines but exhibit high risks for biosafety. Subunit vaccines enjoy high biosafety and specificity but have a few weak points compared to inactivated virus or live attenuated virus vaccines, especially in low immunogenicity. In this study, we developed a new subunit vaccine platform for a potent, adjuvant-free, and multivalent vaccination. The ectodomains of hemagglutinins (HAs) of influenza viruses were expressed in plants as trimers (tHAs) to mimic their native forms. tHAs in plant extracts were directly used without purification for binding to inactivated Lactococcus (iLact) to produce iLact-tHAs, an antigen-carrying bacteria-like particle (BLP). tHAs BLP showed strong immune responses in mice and chickens without adjuvants. Moreover, simultaneous injection of two different antigens by two different formulas, tHAH5N6 + H9N2 BLP or a combination of tHAH5N6 BLP and tHAH9N2 BLP, led to strong immune responses to both antigens. Based on these results, we propose combinations of plant-based antigen production and BLP-based delivery as a highly potent and cost-effective platform for multivalent vaccination for subunit vaccines.
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    Genome-wide profiling of circular RNAs, alternative splicing, and R-loops in stem-differentiating xylem of Populus trichocarpa
    Xuqing Liu, Yubang Gao, Jiakai Liao, Miao Miao, Kai Chen, Feihu Xi, Wentao Wei, Huihui Wang, Yongsheng Wang, Xi Xu, Anireddy S.N. Reddy and Lianfeng Gu
    J Integr Plant Biol 2021, 63 (7): 1294-1308.  
    DOI: 10.1111/jipb.13081
    Abstract (Browse 302)  |   Save
    Circular RNAs (circRNAs) are a recently discovered type of non-coding RNA derived from pre-mRNAs. R-loops consist of a DNA:RNA hybrid and the associated single-stranded DNA. In Arabidopsis thaliana, circRNA:DNA R-loops regulate alternative splicing (AS) of SEPALLATA3 (SEP3). However, the occurrence and functions of circRNAs and R-loops in Populus trichocarpa are largely unexplored. Here, we performed circRNA-enriched sequencing in the stem-differentiating xylem (SDX) of P. trichocarpa and identified 2,742 distinct circRNAs, including circ-CESA4, circ-IRX7, and circ-GUX1, which are generated from genes involved in cellulose, and hemicellulose biosynthesis, respectively. To investigate the roles of circRNAs in modulating alternative splicing (AS), we detected 7,836 AS events using PacBio Iso-Seq and identified 634 circRNAs that overlapped with 699 AS events. Furthermore, using DNA:RNA hybrid immunoprecipitation followed by sequencing (DRIP-seq), we identified 8,932 R-loop peaks that overlapped with 181 circRNAs and 672 AS events. Notably, several SDX-related circRNAs overlapped with R-loop peaks, pointing to their possible roles in modulating AS in SDX. Indeed, overexpressing circ-IRX7 increased the levels of R-loop structures and decreased the frequency of intron retention in linear IRX7 transcripts. This study provides a valuable R-loop atlas resource and uncovers the interplay between circRNAs and AS in SDX of P. trichocarpa.
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    MP3RNA-seq: Massively parallel 3′ end RNA sequencing for high-throughput gene expression profiling and genotyping
    Jian Chen, Xiangbo Zhang, Fei Yi, Xiang Gao, Weibin Song, Haiming Zhao and Jinsheng Lai
    J Integr Plant Biol 2021, 63 (7): 1227-1239.  
    doi: 10.1111/jipb.13077
    Abstract (Browse 318)  |   Save
    Transcriptome deep sequencing (RNA-seq) has become a routine method for global gene expression profiling. However, its application to large-scale experiments remains limited by cost and labor constraints. Here we describe a massively parallel 3′ end RNA-seq (MP3RNA-seq) method that introduces unique sample barcodes during reverse transcription to permit sample pooling immediately following this initial step. MP3RNA-seq allows for handling of hundreds of samples in a single experiment, at a cost of about $6 per sample for library construction and sequencing. MP3RNA-seq is effective for not only high-throughput gene expression profiling, but also genotyping. To demonstrate its utility, we applied MP3RNA-seq to 477 double haploid lines of maize. We identified 19,429 genes expressed in at least 50% of the lines and 35,836 high-quality single nucleotide polymorphisms for genotyping analysis. Armed with these data, we performed expression and agronomic trait quantitative trait locus (QTL) mapping and identified 25,797 expression QTLs for 15,335 genes and 21 QTLs for plant height, ear height, and relative ear height. We conclude that MP3RNA-seq is highly reproducible, accurate, and sensitive for high-throughput gene expression profiling and genotyping, and should be generally applicable to most eukaryotic species.
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    Exploring the diversity of plant proteome
    Yanmei Chen, Yi Wang, Jun Yang, Wenbin Zhou and Shaojun Dai
    J Integr Plant Biol 2021, 63 (7): 1197-1210.  
    DOI: 10.1111/jipb.13087
    Abstract (Browse 434)  |   Save
    The tremendous functional, spatial, and temporal diversity of the plant proteome is regulated by multiple factors that continuously modify protein abundance, modifications, interactions, localization, and activity to meet the dynamic needs of plants. Dissecting the proteome complexity and its underlying genetic variation is attracting increasing research attention. Mass spectrometry (MS)-based proteomics has become a powerful approach in the global study of protein functions and their relationships on a systems level. Here, we review recent breakthroughs and strategies adopted to unravel the diversity of the proteome, with a specific focus on the methods used to analyze posttranslational modifications (PTMs), protein localization, and the organization of proteins into functional modules. We also consider PTM crosstalk and multiple PTMs temporally regulating the life cycle of proteins. Finally, we discuss recent quantitative studies using MS to measure protein turnover rates and examine future directions in the study of the plant proteome.
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    Developing a new model system for potato genetics by androgenesis
    Miru Du, Ting Wang, Qun Lian, Xiaojie Zhang, Guohui Xin, Yuanyuan Pu, Glenn J. Bryan and Jianjian Qi
    J Integr Plant Biol 2021, 63 (4): 628-633.  
    doi: 10.1111/jipb.13018
    Abstract (Browse 405)  |   Save
    High heterozygosity and tetrasomic inheritance complicate studies of asexually propagated polyploids, such as potato. Reverse genetics approaches, especially mutant library construction, can be an ideal choice if a proper mutagenesis genotype is available. Here, we aimed to generate a model system for potato research using anther cultures of Solanum verrucosum, a self‐compatible diploid potato with strong late blight resistance. Six of the 23 regenerants obtained (SVA4, SVA7, SVA22, SVA23, SVA32, and SVA33) were diploids, and their homozygosity was estimated to be >99.99% with 22 polymorphic InDel makers. Two lines—SVA4 and SVA32—had reduced stature (plant height ≤80 cm), high seed yield (>1,000 seeds/plant), and good tuber set (>30 tubers/plant). We further confirmed the full homozygosity of SVA4 and SVA32 using whole‐genome resequencing. These two regenerants possess all the characteristics of a model plant: diploidy, 100% homozygosity, self‐compatibility, and amenability to transgenesis. Thus, we have successfully generated two lines, SVA4 and SVA32, which can potentially be used for mutagenesis and as model plants to rejuvenate current methods of conducting potato research.
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    Multiplex and optimization of dCas9‐TV‐mediated gene activation in plants
    Xiangyu Xiong, Jieping Liang, Zhenxiang Li, Ben‐Qiang Gong and Jian‐Feng Li
    J Integr Plant Biol 2021, 63 (4): 634-645.  
    doi: 10.1111/jipb.13023
    Abstract (Browse 432)  |   Save
    Synthetic gene activators consisting of nuclease‐dead Cas9 (dCas9) for single‐guide RNA (sgRNA)‐directed promoter binding and a transcriptional activation domain (TAD) represent new tools for gene activation from endogenous genomic locus in basic and applied plant research. However, multiplex gene coactivation by dCas9‐TADs has not been demonstrated in whole plants. There is also room to optimize the performance of these tools. Here, we report that our previously developed gene activator, dCas9‐TV, could simultaneously upregulate OsGW7 and OsER1 in rice by up to 3,738 fold, with one sgRNA targeting to each promoter. The gene coactivation could persist to at least the fourth generation. Astonishingly, the polycistronic tRNA‐sgRNA expression under the maize ubiquitin promoter, a Pol II promoter, could cause enormous activation of these genes by up to >40,000‐fold in rice. Moreover, the yeast GCN4 coiled coil‐mediated dCas9‐TV dimerization appeared to be promising for enhancing gene activation. Finally, we successfully introduced a self‐amplification loop for dCas9‐TV expression in Arabidopsis to promote the transcriptional upregulation of AtFLS2, a previously characterized dCas9‐TV‐refractory gene with considerable basal expression. Collectively, this work illustrates the robustness of dCas9‐TV in multigene coactivation and provides broadly useful strategies for boosting transcriptional activation efficacy of dCas9‐TADs in plants.
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    The CBP/p300 histone acetyltransferases function as plant‐specific MEDIATOR subunits in Arabidopsis
    Jing Guo, Long Wei, Shan‐Shan Chen, Xue‐Wei Cai, Yin‐Na Su, Lin Li, She Chen and Xin‐Jian He
    J Integr Plant Biol 2021, 63 (4): 755-771.  
    DOI: 10.1111/jipb.13052
    Abstract (Browse 305)  |   Save
    In eukaryotes, MEDIATOR is a conserved multi‐subunit complex that links transcription factors and RNA polymerase II and that thereby facilitates transcriptional initiation. Although the composition of MEDIATOR has been well studied in yeast and mammals, relatively little is known about the composition of MEDIATOR in plants. By affinity purification followed by mass spectrometry, we identified 28 conserved MEDIATOR subunits in Arabidopsis thaliana, including putative MEDIATOR subunits that were not previously validated. Our results indicated that MED34, MED35, MED36, and MED37 are not Arabidopsis MEDIATOR subunits, as previously proposed. Our results also revealed that two homologous CBP/p300 histone acetyltransferases, HAC1 and HAC5 (HAC1/5) are in fact plant‐specific MEDIATOR subunits. The MEDIATOR subunits MED8 and MED25 (MED8/25) are partially responsible for the association of MEDIATOR with HAC1/5, MED8/25 and HAC1/5 co‐regulate gene expression and thereby affect flowering time and floral development. Our in vitro observations indicated that MED8 and HAC1 form liquid‐like droplets by phase separation, and our in vivo observations indicated that these droplets co‐localize in the nuclear bodies at a subset of nuclei. The formation of liquid‐like droplets is required for MED8 to interact with RNA polymerase II. In summary, we have identified all of the components of Arabidopsis MEDIATOR and revealed the mechanism underlying the link of histone acetylation and transcriptional regulation.
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    Comparative transcriptomics reveals hidden issues in the plant response to arthropod herbivores
    M. Estrella Santamaria, Alejandro Garcia, Ana Arnaiz, Irene Rosa‐Diaz, Gara Romero‐Hernandez, IsabelDiaz and Manuel Martinez
    J Integr Plant Biol 2021, 63 (2): 312-326.  
    doi: 10.1111/jipb.13026
    Abstract (Browse 330)  |   Save
    Plants experience different abiotic/biotic stresses, which trigger their molecular machinery to cope with them. Besides general mechanisms prompted by many stresses, specific mechanisms have been introduced to optimize the response to individual threats. However, these key mechanisms are difficult to identify. Here, we introduce an in‐depth species‐specific transcriptomic analysis and conduct an extensive meta‐analysis of the responses to related species to gain more knowledge about plant responses. The spider mite Tetranychus urticae was used as the individual species, several arthropod herbivores as the related species for meta‐analysis, and Arabidopsis thaliana plants as the common host. The analysis of the transcriptomic data showed typical common responses to herbivory, such as jasmonate signaling or glucosinolate biosynthesis. Also, a specific set of genes likely involved in the particularities of the Arabidopsis‐spider mite interaction was discovered. The new findings have determined a prominent role in this interaction of the jasmonate‐induced pathways leading to the biosynthesis of anthocyanins and tocopherols. Therefore, tandem individual/general transcriptomic profiling has been revealed as an effective method to identify novel relevant processes and specificities in the plant response to environmental stresses.
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    Three‐dimensional reconstruction and comparison of vacuolar membranes in response to viral infection
    Xueting Wang, Juncai Ma, Xuejiao Jin, Ning Yue, Peng Gao, Keith Ka Ki Mai, Xian‐Bing Wang, Dawei Li, Byung‐Ho Kang and Yongliang Zhang
    J Integr Plant Biol 2021, 63 (2): 353-364.  
    doi: 10.1111/jipb.13027
    Abstract (Browse 306)  |   Save
    The vacuole is a unique plant organelle that plays an important role in maintaining cellular homeostasis under various environmental stress conditions. However, the effects of biotic stress on vacuole structure has not been examined using three‐dimensional (3D) visualization. Here, we performed 3D electron tomography to compare the ultrastructural changes in the vacuole during infection with different viruses. The 3D models revealed that vacuoles are remodeled in cells infected with cucumber mosaic virus (CMV) or tobacco necrosis virus A Chinese isolate (TNV‐AC), resulting in the formation of spherules at the periphery of the vacuole. These spherules contain neck‐like channels that connect their interior with the cytosol. Confocal microscopy of CMV replication proteins 1a and 2a and TNV‐AC auxiliary replication protein p23 showed that all of these proteins localize to the tonoplast. Electron microscopy revealed that the expression of these replication proteins alone is sufficient to induce spherule formation on the tonoplast, suggesting that these proteins play prominent roles in inducing vacuolar membrane remodeling. This is the first report of the 3D structures of viral replication factories built on the tonoplasts. These findings contribute to our understanding of vacuole biogenesis under normal conditions and during assembly of plant (+) RNA virus replication complexes.
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    Benefiting others and self: Production of vitamins in plants
    Yufei Li, Chenkun Yang, Hasan Ahmad, Mohamed Maher, Chuanying Fang and Jie Luo
    J Integr Plant Biol 2021, 63 (1): 210-227.  
    doi: 10.1111/jipb.13047
    Abstract (Browse 288)  |   Save
    Vitamins maintain growth and development in humans, animals, and plants. Because plants serve as essential producers of vitamins, increasing the vitamin contents in plants has become a goal of crop breeding worldwide. Here, we begin with a summary of the functions of vitamins. We then review the achievements to date in elucidating the molecular mechanisms underlying how vitamins are synthesized, transported, and regulated in plants. We also stress the exploration of variation in vitamins by the use of forward genetic approaches, such as quantitative trait locus mapping and genome‐wide association studies. Overall, we conclude that exploring the diversity of vitamins could provide new insights into plant metabolism and crop breeding.
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    The plant cell wall: Biosynthesis, construction, and functions
    Baocai Zhang, Yihong Gao, Lanjun Zhang and Yihua Zhou
    J Integr Plant Biol 2021, 63 (1): 251-272.  
    doi: 10.1111/jipb.13055
    Abstract (Browse 511)  |   Save
    The plant cell wall is composed of multiple biopolymers, representing one of the most complex structural networks in nature. Hundreds of genes are involved in building such a natural masterpiece. However, the plant cell wall is the least understood cellular structure in plants. Due to great progress in plant functional genomics, many achievements have been made in uncovering cell wall biosynthesis, assembly, and architecture, as well as cell wall regulation and signaling. Such information has significantly advanced our understanding of the roles of the cell wall in many biological and physiological processes and has enhanced our utilization of cell wall materials. The use of cutting‐edge technologies such as single‐molecule imaging, nuclear magnetic resonance spectroscopy, and atomic force microscopy has provided much insight into the plant cell wall as an intricate nanoscale network, opening up unprecedented possibilities for cell wall research. In this review, we summarize the major advances made in understanding the cell wall in this era of functional genomics, including the latest findings on the biosynthesis, construction, and functions of the cell wall.
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