园艺-分子生物合辑Horticulture — Genetics · Breeding
|Comparative transcriptome analysis provides insights into the mechanism of pear dwarfing
|TANG Zi-kai1*, SUN Man-yi1*, LI Jia-ming1, SONG Bo-bo1, LIU Yue-yuan1, TIAN Yi-ke2, WANG Cai-hong2, WU Jun1
|1 College of Horticulture, State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, Nanjing 210095, P.R.China
2 College of Horticulture, Qingdao Agricultural University, Qingdao 266109, P.R.China
本研究以‘矮玉香’和‘翠冠’为父母本，通过杂交获得矮化性状的后代（DPP）和乔化表型的后代（APP）。与APP相比，DPP植株高度降低了62.8%，节间长度显著缩短。茎尖组织切片观察显示，DPP细胞形状不规则，细胞平均长度较大，但纵向排布的细胞数量显著减少。两种表型梨之间的交叉嫁接试验表明，‘矮生梨’的矮化表型主要由地上部分决定，与根系无关。根据二者茎尖的RNAseq数据共鉴定出1401个差异表达基因,其中包含101转录因子。功能富集分析显示，植物激素、细胞分裂、细胞壁代谢等相关通路中的基因显著差异表达。在DPP中，若干BR信号转导和细胞周期相关基因表达显著下调；同时，一些调控BR和GA降解的基因表达上调。结合切片观察的结果与RNA-seq的数据推断，DPP的矮化表型主要归因于细胞分裂的不足。为进一步缩小候选基因集，将差异表达基因映射到前人所做的‘Le Nain Vert’矮化性状定位区间，共鉴定出4个关键基因。结合功能注释分析，发现其中一个DELLA基因可能在调控‘矮生梨’矮化表型中发挥重要的作用。本研究结果为进一步探索梨矮化的遗传和分子机制奠定了基础。
Abstract Dwarfism is an important trait which is closely related to the efficiency of fruit orchard management and production. However, dwarfing cannot be widely applied in the cultivation of pears, especially Asian pears. Developing varieties with dwarf characteristics is a goal of paramount importance in pear breeding. In the present study, dwarf phenotype pears (DPPs) and arborescent phenotype pears (APPs) were obtained from the offspring of a cross between ‘Aiyuxiang’ and ‘Cuiguan’ pear cultivars, which exhibited dwarfed and arborescent statures, respectively. When compared with APPs, the heights of DPPs showed a 62.8% reduction, and the internode lengths were significantly shorter. Cross-grafting between DPPs and APPs demonstrated that the dwarfed phenotype of DPPs was primarily induced by the aerial portions of the plant, and independent of the root system. Observations of stem tissue sections showed that DPP cells were arranged chaotically with irregular shapes, and the average length was larger than that of the APP cells. A total of 1 401 differently expressed genes (DEGs) in shoot apices between DPPs and APPs were identified by RNA-sequencing (RNA-Seq), and these DEGs were mainly enriched in the ‘phytohormone-related pathways, cell wall metabolism and cell division’ categories. Moreover, 101 DEGs were identified as transcription factors (TFs). In DPPs, several brassinosteroids (BR) signaling and cell cycle-related genes were significantly down-regulated, while genes involved in BR and GA degradation were up-regulated. Comprehensive analysis of RNA-Seq data and stem tissue sections suggested that the dwarfed phenotype of DPPs could be primarily attributed to deficiencies in cell division. Previous work using simple sequence repeat (SSR) markers narrowed the location of the gene responsible for the dwarf phenotype of ‘Le Nain Vert’. Through combined analysis of our transcriptomic data with the SSR results, we identified four genes as promising candidates for the dwarf phenotype, among which, a DELLA gene could be the most promising. The results presented in this study provide a sound foundation for further exploration into the genetic and molecular mechanisms underlying pear dwarfing.
Received: 21 March 2021
Accepted: 10 June 2021
|Fund: This work was supported by the National Key Research and Development Program of China (2018YFD1000200), the Earmarked Fund for Jiangsu Agricultural Industry Technology System, China (JATS 401), the National Science Foundation of China (31801835), the China Agriculture Research System of MOF and MARA (CARS-28), the Natural Science Foundation of Jiangsu Province, China (BK20180516), and the Agricultural Variety Improvement Project of Shandong Province, China (2019LZGC008).
|About author: TANG Zi-kai, E-mail: email@example.com; SUN Man-yi, E-mail: firstname.lastname@example.org; Correspondence WU Jun, E-mail: email@example.com; WANG Cai-hong, E-mail: firstname.lastname@example.org
* These authors contributed equally to this study.
Cite this article:
TANG Zi-kai, SUN Man-yi, LI Jia-ming, SONG Bo-bo, LIU Yue-yuan, TIAN Yi-ke, WANG Cai-hong, WU Jun.
Comparative transcriptome analysis provides insights into the mechanism of pear dwarfing. Journal of Integrative Agriculture, 21(7): 1952-1967.
| Achard P, Gong F, Cheminant S, Alioua M, Hedden P, Genschik P. 2008. The cold-inducible CBF1 factor-dependent signaling pathway modulates the accumulation of the growth-repressing DELLA proteins via its effect on gibberellin metabolism. The Plant Cell, 20, 2117–2129.
Bolger A M, Lohse M, Usadel B. 2014. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics, 30, 2114–2120.
Cantin C M, Arus P, Eduardo I. 2018. Identification of a new allele of the Dw gene causing brachytic dwarfing in peach. BMC Research Notes, 11, 386.
Chen J Y, Dai X F. 2010. Cloning and characterization of the Gossypium hirsutum major latex protein gene and functional analysis in Arabidopsis thaliana. Planta, 231, 861–873.
Chen L, Yang H, Fang Y, Guo W, Chen H, Zhang X, Dai W, Chen S, Hao Q, Yuan S, Zhang C, Huang Y, Shan Z, Yang Z, Qiu D, Liu X, Tran L P, Zhou X, Cao D. 2020. Overexpression of GmMYB14 improves high-density yield and drought tolerance of soybean through regulating plant architecture mediated by the brassinosteroid pathway. Plant Biotechnology Journal, 19, 702–716.
Chen L G, Gao Z, Zhao Z, Liu X, Li Y, Zhang Y, Liu X, Sun Y, Tang W. 2019. BZR1 family transcription factors function redundantly and indispensably in BR signaling but exhibit BRI1-independent function in regulating anther development in Arabidopsis. Molecular Plant, 12, 1408–1415.
Chen Z, Zhao P X, Miao Z Q, Qi G F, Wang Z, Yuan Y, Ahmad N, Cao M J, Hell R, Wirtz M, Xiang C B. 2019. SULTR3s function in chloroplast sulfate uptake and affect ABA biosynthesis and the stress response. Plant Physiology, 180, 593–604.
Cheng J, Zhang M, Tan B, Jiang Y, Zheng X, Ye X, Guo Z, Xiong T, Wang W, Li J, Feng J. 2019. A single nucleotide mutation in GID1c disrupts its interaction with DELLA1 and causes a GA-insensitive dwarf phenotype in peach. Plant Biotechnology Journal, 17, 1723–1735.
Deepika, Ankit, Sagar S, Singh A. 2020. Dark-induced hormonal regulation of plant growth and development. Frontiers in Plant Science, 11, 581666.
Dill A, Jung H S, Sun T P. 2001. The DELLA motif is essential for gibberellin-induced degradation of RGA. Proceedings of the National Academy of Sciences of the United States of America, 98, 14162–14167.
Fabian T, Lorbiecke R, Umeda M, Sauter M. 2000. The cell cycle genes cycA1;1 and cdc2Os-3 are coordinately regulated by gibberellin in planta. Planta, 211, 376–383.
Feng Y, Yin Y, Fei S. 2015. Down-regulation of BdBRI1, a putative brassinosteroid receptor gene produces a dwarf phenotype with enhanced drought tolerance in Brachypodium distachyon. Plant Science, 234, 163–173.
Fu F Q, Mao W H, Shi K, Zhou Y H, Asami T, Yu J Q. 2008. A role of brassinosteroids in early fruit development in cucumber. Journal of Experimental Botany, 59, 2299–2308.
Gan Z, Wang Y, Wu T, Xu X, Zhang X, Han Z. 2018. MdPIN1b encodes a putative auxin efflux carrier and has different expression patterns in BC and M9 apple rootstocks. Plant Molecular Biology, 96, 353–365.
Guo M, Simmons C R. 2011. Cell number counts - the fw2.2 and CNR genes and implications for controlling plant fruit and organ size. Plant Science, 181, 1–7.
Han H, Liu X, Zhou Y. 2020. Transcriptional circuits in control of shoot stem cell homeostasis. Current Opinion in Plant Biology, 53, 50–56.
Hollender C A, Hadiarto T, Srinivasan C, Scorza R, Dardick C. 2016. A brachytic dwarfism trait (dw) in peach trees is caused by a nonsense mutation within the gibberellic acid receptor PpeGID1c. New Phytologist, 210, 227–239.
Hu Y X, Tao Y B, Xu Z F. 2017. Overexpression of Jatropha Gibberellin 2-oxidase 6 (JcGA2ox6) induces dwarfism and smaller leaves, flowers and fruits in Arabidopsis and Jatropha. Frontiers in Plant Science, 8, 2103.
Jones A R, Forero-Vargas M, Withers S P, Smith R S, Traas J, Dewitte W, Murray J A H. 2017. Cell-size dependent progression of the cell cycle creates homeostasis and flexibility of plant cell size. Nature Communications, 8, 15060.
Kim D, Paggi J M, Park C, Bennett C, Salzberg S L. 2019. Graph-based genome alignment and genotyping with HISAT2 and HISAT-genotype. Nature Biotechnology, 37, 907–915.
Liao Y, Smyth G K, Shi W. 2014. featureCounts: an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics, 30, 923–930.
Liu K, Li Y, Chen X, Li L, Liu K, Zhao H, Wang Y, Han S. 2018. ERF72 interacts with ARF6 and BZR1 to regulate hypocotyl elongation in Arabidopsis. Journal of Experimental Botany, 69, 3933–3947.
Lu Y, Feng Z, Meng Y, Bian L, Xie H, Mysore K S, Liang J. 2020. SLENDER RICE1 and Oryza sativa INDETERMINATE DOMAIN2 regulating OsmiR396 are involved in stem elongation. Plant Physiology, 182, 2213–2227.
Ma Z, Wu T, Huang K, Jin Y M, Li Z, Chen M, Yun S, Zhang H, Yang X, Chen H, Bai H, Du L, Ju S, Guo L, Bian M, Hu L, Du X, Jiang W. 2020. A novel AP2/ERF transcription factor, OsRPH1, negatively regulates plant height in rice. Frontiers in Plant Science, 11, 709.
Nemoto K, Ramadan A, Arimura G I, Imai K, Tomii K, Shinozaki K, Sawasaki T. 2017. Tyrosine phosphorylation of the GARU E3 ubiquitin ligase promotes gibberellin signalling by preventing GID1 degradation. Nature Communications, 8, 1004.
Nguyen H P, Jeong H Y, Jeon S H, Kim D, Lee C. 2017. Rice pectin methylesterase inhibitor28 (OsPMEI28) encodes a functional PMEI and its overexpression results in a dwarf phenotype through increased pectin methylesterification levels. Journal of Plant Physiology, 208, 17–25.
Niu M, Wang Y, Wang C, Lyu J, Wang Y, Dong H, Long W, Wang D, Kong W, Wang L, Guo X, Sun L, Hu T, Zhai H, Wang H, Wan J. 2017. ALR encoding dCMP deaminase is critical for DNA damage repair, cell cycle progression and plant development in rice. Journal of Experimental Botany, 68, 5773–5786.
Nomura T, Magome H, Hanada A, Takeda-Kamiya N, Mander L N, Kamiya Y, Yamaguchi S. 2013. Functional analysis of Arabidopsis CYP714A1 and CYP714A2 reveals that they are distinct gibberellin modification enzymes. Plant & Cell Physiology, 54, 1837–1851.
Oh E, Zhu J Y, Ryu H, Hwang I, Wang Z Y. 2014. TOPLESS mediates brassinosteroid-induced transcriptional repression through interaction with BZR1. Nature Communications, 5, 4140.
Ohnishi T, Godza B, Watanabe B, Fujioka S, Hategan L, Ide K, Shibata K, Yokota T, Szekeres M, Mizutani M. 2012. CYP90A1/CPD, a brassinosteroid biosynthetic cytochrome P450 of Arabidopsis, catalyzes C-3 oxidation. Journal of Biological Chemistry, 287, 31551–31560.
Ou C, Wang F, Wang J, Li S, Zhang Y, Fang M, Ma L, Zhao Y, Jiang S. 2019. A de novo genome assembly of the dwarfing pear rootstock Zhongai 1. Scientific Data, 6, 281.
Pang H, Yan Q, Zhao S, He F, Xu J, Qi B, Zhang Y. 2019. Knockout of the S-acyltransferase gene, PbPAT14, confers the dwarf yellowing phenotype in first generation pear by ABA accumulation. International Journal of Molecular Sciences, 20, 6347.
Qi L, Chen L, Wang C, Zhang S, Yang Y, Liu J, Li D, Song J, Wang R. 2020. Characterization of the auxin efflux transporter PIN proteins in pear. Plants (Basel), 9, 349.
Qi W, Sun F, Wang Q, Chen M, Huang Y, Feng Y Q, Luo X, Yang J. 2011. Rice ethylene-response AP2/ERF factor OsEATB restricts internode elongation by down-regulating a gibberellin biosynthetic gene. Plant Physiology, 157, 216–228.
Ripetti V, Escoute J, Verdeil J L, Costes E. 2008. Shaping the shoot: the relative contribution of cell number and cell shape to variations in internode length between parent and hybrid apple trees. Journal of Experimental Botany, 59, 1399–1407.
Sakamoto T, Kawabe A, Tokida-Segawa A, Shimizu B, Takatsuto S, Shimada Y, Fujioka S, Mizutani M. 2011. Rice CYP734As function as multisubstrate and multifunctional enzymes in brassinosteroid catabolism. Plant Journal, 67, 1–12.
Sazuka T, Kamiya N, Nishimura T, Ohmae K, Sato Y, Imamura K, Nagato Y, Koshiba T, Nagamura Y, Ashikari M, Kitano H, Matsuoka M. 2009. A rice tryptophan deficient dwarf mutant, tdd1, contains a reduced level of indole acetic acid and develops abnormal flowers and organless embryos. Plant Journal, 60, 227–241.
Serrano-Mislata A, Bencivenga S, Bush M, Schiessl K, Boden S, Sablowski R. 2017. DELLA genes restrict inflorescence meristem function independently of plant height. Nature Plants, 3, 749–754.
Speicher T L, Li P Z, Wallace I S. 2018. Phosphoregulation of the plant cellulose synthase complex and cellulose synthase-like proteins. Plants (Basel), 7, 52.
Sun S, Chen D, Li X, Qiao S, Shi C, Li C, Shen H, Wang X. 2015. Brassinosteroid signaling regulates leaf erectness in Oryza sativa via the control of a specific U-type cyclin and cell proliferation. Developmental Cell, 34, 220–228.
Takahashi N, Nakazawa M, Shibata K, Yokota T, Ishikawa A, Suzuki K, Kawashima M, Ichikawa T, Shimada H, Matsui M. 2005. shk1-D, a dwarf Arabidopsis mutant caused by activation of the CYP72C1 gene, has altered brassinosteroid levels. Plant Journal, 42, 13–22.
Takatsuka H, Umeda-Hara C, Umeda M. 2015. Cyclin-dependent kinase-activating kinases CDKD;1 and CDKD;3 are essential for preserving mitotic activity in Arabidopsis thaliana. Plant Journal, 82, 1004–1017.
Tang H, Wang X, Bowers J E, Ming R, Alam M, Paterson A H. 2008. Unraveling ancient hexaploidy through multiply-aligned angiosperm gene maps. Genome Research, 18, 1944–1954.
Tao Y, Miao J, Wang J, Li W, Xu Y, Wang F, Jiang Y, Chen Z, Fan F, Xu M, Zhou Y, Liang G, Yang J. 2020. RGG1, involved in the cytokinin regulatory pathway, controls grain size in rice. Rice (N Y), 13, 76.
Unterholzner S J, Rozhon W, Papacek M, Ciomas J, Lange T, Kugler K G, Mayer K F, Sieberer T, Poppenberger B. 2015. Brassinosteroids are master regulators of gibberellin biosynthesis in Arabidopsis. The Plant Cell, 27, 2261–2272.
Vilarrasa-Blasi J, González-García M P, Frigola D, Fàbregas-Vallvé N, Alexiou K G, López-Bigas N, Rivas S, Jauneau A, Lohmann J U, Benfey P N, Ibañes M, Caño-Delgado A I. 2015. Regulation of plant stem cell quiescence by a brassinosteroid signaling module. Developmental Cell, 30, 36–47.
Wang C, Tian Y, Buck E J, Gardiner S E, Dai H, Jia Y. 2011. Genetic mapping of PcDw determining pear dwarf trait. Journal of the American Society for Horticultural Science, 136, 48–53.
Wang C H, Li W, Tian Y K, Hou D L, Bai M D. 2016. Development of molecular markers for genetic and physical mapping of the PcDw locus in pear (Pyrus communis L.). The Journal of Horticultural Science and Biotechnology, 91, 299–307.
Wang J, Jiang J, Wang J, Chen L, Fan S L, Wu J W, Wang X, Wang Z X. 2014. Structural insights into the negative regulation of BRI1 signaling by BRI1-interacting protein BKI1. Cell Research, 24, 1328–1341.
Wang S S, Liu Z Z, Sun C, Shi Q H, Yao Y X, You C X, Hao Y J. 2012. Functional characterization of the apple MhGAI1 gene through ectopic expression and grafting experiments in tomatoes. Journal of Plant Physiology, 169, 303–310.
Wang Y, Tang H, Debarry J D, Tan X, Li J, Wang X, Lee T H, Jin H, Marler B, Guo H, Kissinger J C, Paterson A H. 2012. MCScanX: a toolkit for detection and evolutionary analysis of gene synteny and collinearity. Nucleic Acids Research, 40, e49.
Wu J, Wang Z, Shi Z, Zhang S, Ming R, Zhu S, Khan M A, Tao S, Korban S S, Wang H, Chen N J, Nishio T, Xu X, Cong L, Qi K, Huang X, Wang Y, Zhao X, Wu J, Deng C, et al. 2013. The genome of the pear (Pyrus bretschneideri Rehd.). Genome Research, 23, 396–408.
Xiao K, Chen W, Chen X, Zhu X, Guan P, Hu J. 2020. CCS52 and DEL1 function in root-knot nematode giant cell development in Xinjiang wild myrobalan plum (Prunus sogdiana Vassilcz). Protoplasma, 257, 1333–1344.
Xiao Y, Liu D, Zhang G, Tong H, Chu C. 2017. Brassinosteroids regulate OFP1, a DLT interacting protein, to modulate plant architecture and grain morphology in rice. Frontiers in Plant Science, 8, 1698.
Xiao Y X, Wang C H, Tian Y K, Yang S L, Shen J L, Liu Q Q, Zhang H Y. 2019. Candidates responsible for dwarf pear phenotype as revealed by comparative transcriptome analysis. Molecular Breeding, 39, 10.
Xing Y, Wang N, Zhang T, Zhang Q, Du D, Chen X, Lu X, Zhang Y, Zhu M, Liu M, Sang X, Li Y, Ling Y, He G. 2020. SHORT-ROOT 1 is critical to cell division and tracheary element development in rice roots. Plant Journal, 105, 1179–1191.
Xu X, Du X, Wang F, Sha J, Chen Q, Tian G, Zhu Z, Ge S, Jiang Y. 2020. Effects of potassium levels on plant growth, accumulation and distribution of carbon, and nitrate metabolism in apple dwarf rootstock seedlings. Frontiers in Plant Science, 11, 904.
Yan F, Hu G, Ren Z, Deng W, Li Z. 2015. Ectopic expression a tomato KNOX Gene Tkn4 affects the formation and the differentiation of meristems and vasculature. Plant Molecular Biology, 89, 589–605.
Zentella R, Sui N, Barnhill B, Hsieh W P, Hu J, Shabanowitz J, Boyce M, Olszewski N E, Zhou P, Hunt D F, Sun T P. 2017. The Arabidopsis O-fucosyltransferase SPINDLY activates nuclear growth repressor DELLA. Nature Chemical Biology, 13, 479–485.
Zheng X, Zhao Y, Shan D, Shi K, Wang L, Li Q, Wang N, Zhou J, Yao J, Xue Y, Fang S, Chu J, Guo Y, Kong J. 2018. MdWRKY9 overexpression confers intensive dwarfing in the M26 rootstock of apple by directly inhibiting brassinosteroid synthetase MdDWF4 expression. New Phytologist, 217, 1086–1098.
Zheng Y, Jiao C, Sun H, Rosli H G, Pombo M A, Zhang P, Banf M, Dai X, Martin G B, Giovannoni J J, Zhao P X, Rhee S Y, Fei Z. 2016. iTAK: A program for genome-wide prediction and classification of plant transcription factors, transcriptional regulators, and protein kinases. Molecular Plant, 9, 1667–1670.
Zhou X, Zhang Z L, Park J, Tyler L, Yusuke J, Qiu K, Nam E A, Lumba S, Desveaux D, McCourt P, Kamiya Y, Sun T P. 2016. The ERF11 transcription factor promotes internode elongation by activating gibberellin biosynthesis and signaling. Plant Physiology, 171, 2760–2670.
|No Suggested Reading articles found!