Cloning and Spatiotemporal Expression Analysis of PgWUS and PgBEL1 in Punica granatum

doi:10.16420/j.issn.0513-353x.2020-0377

Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (2): 355-366.doi: 10.16420/j.issn.0513-353x.2020-0377

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Cloning and Spatiotemporal Expression Analysis of PgWUS and PgBEL1 in Punica granatum

ZHAO Yujie, LIU Cuiyu, ZHAO Xueqing, WANG Yuying, YAN Ming, YUAN Zhaohe^*()

College of Forestry,Nanjing Forestry University,Nanjing 210037,China

Received:2020-08-04 Revised:2020-11-01 Online:2021-02-25 Published:2021-03-09
Contact: YUAN Zhaohe E-mail:zhyuan88@hotmail.com

Abstract

Abstract:

Based on the pomegranate genome database,PgWUS and PgBEL1,the homolog of WUS and BEL1 were identified and cloned by homologous cloning. The full-length CDS sequences of PgBEL1 and PgWUS were 1 851 bp and 936 bp,encoding 616 and 311 amino acids,respectively. The protein sequence multiple alignment and phylogenetic analysis suggested that PgWUS had the highest evolution relationship with WUS in Cucumis melo var. makuwa and C. sativus. There was high similarity of amino acid sequences in PgBEL1 and BEL1 of Eucatyptus grandis. Subcellular localization results showed that both PgBEL1 and PgWUS were located in the nucleus. Real-time quantitative PCR analysis suggested that the expression level of PgBEL1 in bisexual flowers was higher than that in functional male flowers at P1 P4 periods（bud vertical diameter < 12.0 mm）. PgBEL1 was highly expressed in leaf,it was 3.2 times than that in calyx while 1.2 times than that in stem,respectively. PgBEL1 expression was low in shoot apex. The expression level of PgBEL1 in pistil was 1.16 times than that in stamen. At the P2 and P3 periods of pomegranate flower development that bud vertical diameter was 5.1-10.0 mm,the expression level of PgWUS in bisexual flowers was higher than that in functional male flowers. PgWUS expression was the lowest in calyx and was the highest in stem. The expression level of PgWUS in pistil was 1.5 times than that in stamen and 1.8 times than that in shoot apex. There was a significant difference in the expression levels of PgWUS in pistil and stamen.

Key words: pomegranate, WUS, BEL1, gene clone, expression analysis

CLC Number:

S665.4

ZHAO Yujie, LIU Cuiyu, ZHAO Xueqing, WANG Yuying, YAN Ming, YUAN Zhaohe. Cloning and Spatiotemporal Expression Analysis of PgWUS and PgBEL1 in Punica granatum[J]. Acta Horticulturae Sinica, 2021, 48(2): 355-366.

Figures/Tables 7

Fig. 1 Bisexual flower and functional male flower at eight developmental stages Buds were divided into eight developmental stages according to vertical diameter,3.0-5.0 mm（P1）,5.1-8.0 mm（P2）,8.1-10.0 mm（P3）,10.1-12.0 mm（P4）,12.1-14.0 mm（P5）,14.1-16.0 mm（P6）,16.1-18.0 mm（P7）,18.1-20.0 mm（P8）.

Table 1 Primers for the gene cloning and qRT-PCR

引物 Primer	引物序列（5′-3′） Primer sequence	注释 Annotation
PgBEL1	F：ATGGTCAGAGAACTCTGT;R：TTAACCGGCCAAATCATG	Gene clone
PgWUS	F：ATGGAACAACCTCAGCAGC;R：CTAGGGCGAACTCGGGGAC	Gene clone
qRTPgBEL1	F：CCGACTAAGCCGAAGACC;R：TCGCAGTAATGCCTGTATCT	Gene expression
qRTPgWUS	F：CCAACTCTTTCGGTCCCT;R：TCCATAGTTAGCCATCTGTCC	Gene expression
PgActin	F：AGTCCTCTTCCAGCCATCTC;R：CACTGAGCACAATGTTTCCA	Gene expression

Fig. 2 Phylogenetic tree of pomegranate based on amino acid sequences of WOX（A）and TALE（B）

Fig. 3 Gene cloning of PgWUS and PgBEL1

Fig. 4 Phylogenetic tree of different species based on amino acid of WUS（A）and BEL1（B）

Fig. 5 Relative expression of PgWUS and PgBEL1 at the development of pomegranate flowers Different lowercase letters represent significant difference at 0.05. The same below.

Fig. 6 Relative expression of PgWUS and PgBEL1 in different organs of pomegranate

References 55

[1]	Bao Zhi-ru. 2018. Regeneration of cotyledon explants and functional analysis of ANT and BEL1 genes in Platanus acerifolia[Ph. D. Dissertation]. Wuhan:Huazhong Agricultural University. (in Chinese)
	暴志茹. 2018. 二球悬铃木子叶再生体系的建立及ANT和BEL1同源基因的功能研究[博士论文]. 武汉:华中农业大学.
[2]	Becker A, Bey M, Bürglin T R, Saedler H, Theissen G. 2002. Ancestry and diversity of BEL1-like homeobox genes revealed by gymnosperm (Gnetum gnemon)homologs. Development Genes and Evolution, 212:452-457. doi: 10.1007/s00427-002-0259-7 URL
[3]	Bencivenga S, Simonini S, Benková E, Colombo L. 2012. The transcription factors BEL1 and SPL are required for cytokinin and auxin signaling during ovule development in Arabidopsis. The Plant Cell, 24(7):2886-2897. doi: 10.1105/tpc.112.100164 pmid: 22786869
[4]	Brambilla V, Battaglia R, Colombo M, Masiero S, Bencivenga S, Kater M M, Colombo L. 2007. Genetic and molecular interactions between BELL1 and MADS-box factors support ovule development in Arabidopsis. The Plant Cell, 19(8):2544-56. doi: 10.1105/tpc.107.051797 URL
[5]	Brambilla V, Kater M, Colombo L. 2008. Ovule integument identity determination in Arabidopsis. Plant Signaling Behavior, 3(4):246-247. doi: 10.4161/psb.3.4.5175 URL
[6]	Brown R H, Nickrent D L and Gasser C S. 2010. Expression of ovule and integument-associated genes in reduced ovules of Santalales. Evolution Development, 12(2):231-240. doi: 10.1111/j.1525-142X.2010.00407.x URL
[7]	Byrne M E, Groover A T, Fontana J R, Martienssen R A. 2003. Phyllotactic pattern and stem cell fate are determined by the Arabidopsis homeobox gene BELLRINGER. Development, 130(17):3941-3950. pmid: 12874117
[8]	Cai Yong-li, Lu Xin-gu, Zhu Li-wu. 1993. Preliminary research on flower bud differentiation of‘Pink’pomegranate. Acta Horticulturae Sinica, 20(1):23-26. (in Chinese)
	蔡永立, 卢心固, 朱立武. 1993. ‘粉皮’石榴花芽分化研究. 园艺学报, 20(1):23-26.
[9]	Chen L N, Zhang J, Li H X, Niu J, Xue H, Liu B B, Wang Q, Luo X, Zhang F H, Zhao D G, Cao S Y. 2017. Transcriptomic analysis reveals candidate genes for female sterility in pomegranate flowers. Front Plant science, 8:1430. doi: 10.3389/fpls.2017.01430 URL
[10]	Chen Li-na. 2017. Exploring regulatory genes of pomegranate(Punica granatum)female sterility based on transcriptome analysis[M. D. Dissertation]. Zhengzhou:Chinese Academy of Agricultural Sciences. (in Chinese)
	陈利娜. 2017. 基于转录组测序探讨石榴花雌蕊败育的相关调控基因[硕士论文]. 郑州:中国农业科学院.
[11]	Chen H, Rosin F M, Prat S, Hannapel D J. 2003. Interacting transcription factors from the three-amino acid loop extension superclass regulate tuber formation. Plant Physiology, 132(3):1391-1404. doi: 10.1104/pp.103.022434 URL
[12]	Cho S H, Kang K, Lee S H, Lee I J, Paek N C. 2016. OsWOX3A is involved in negative feedback regulation of the gibberellic acid biosynthetic pathway in rice(Oryza sativa). Journal of Experimental Botany, 67(6):1677-1687. doi: 10.1093/jxb/erv559 URL
[13]	Chou K C, Shen H B. 2010. Plant-mPLoc:a top-down strategy to augment the power for predicting plant protein subcellular localization. PLoS ONE, 5:e11335. doi: 10.1371/journal.pone.0011335 URL
[14]	Deyhle F, Sarkar A K, Tucker E J, Laux T. 2007. WUSCHEL regulates cell differentiation during anther development. Developmental Biology, 302(1):154-159. doi: 10.1016/j.ydbio.2006.09.013 URL
[15]	Groβ-Hardt R, Lenhard M, Laux T. 2002. WUSCHEL signaling functions in interregional communication during Arabidopsis ovule development. Genes Development, 16:1129-1138. doi: 10.1101/gad.225202 URL
[16]	Haecker A, Grob-Hardt R, Geiges B, Sarkar A, Breuninger H, Herrmann M, Laux T. 2004. Expression dynamics of WOX genes mark cell fate decisions during early embryonic patterning in Arabidopsis thaliana. Development, 131(3):657-668. pmid: 14711878
[17]	Huang Z G, Shi T, Zheng B L, Yumul R E, Liu X G, You C J, Gao Z H, Xiao L T, Chen X M. 2017. APETALA2 antagonizes the transcriptional activity of AGAMOUS in regulating floral stem cells in Arabidopsis thaliana. New Phytologist, 215(3):1197-1209. doi: 10.1111/nph.2017.215.issue-3 URL
[18]	Kavai-ool U N, Karpenko O Y, Ezhova T A. 2011. Interaction between the PINOID/ABRUPTUS gene with the AGAMOUS gene negatively regulating stem cells proliferetion in the Arabidopsis thaliana floral meristem. Russian Journal of Developmental Biology, 42(2):120-124. doi: 10.1134/S1062360411020081 URL
[19]	Kieffer M, Stern Y, Cook H, Clerici E, Maulbetsch C, Laux T, Davies B. 2006. Analysis of the transcription factor WUSCHEL and its functional homologue in Antirrhinum reveals a potential mechanism for their roles in meristem maintenance. The Plant Cell, 18(3):560-573. doi: 10.1105/tpc.105.039107 URL
[20]	Kumar R, Kushalappa K, Godt D, Pidkowich M S, Pastorelli S, Hepworth S R, Haughn G W. 2007. The Arabidopsis BEL1-LIKE HOMEODOMAIN proteins SAW1 and SAW2 act redundantly to regulate KNOX expression spatially in leaf margins. The Plant Cell, 19(9):2719-35. doi: 10.1105/tpc.106.048769 URL
[21]	Kumar S, Stecher G, Tamura K. 2016. MEGA7:Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Molecular Biology and Evolution, 33(7):1870-1874. doi: 10.1093/molbev/msw054 URL
[22]	Lenhard M, Bohnert A, Jürgens G, Laux T. 2001. Termination of stem cell maintenance in Arabidopsis floral meristems by interactions between WUSCHEL and AGAMOUS. Cell, 105(6):805-814. pmid: 11440722
[23]	Liu Y, You S, Taylor-Teeples M, Li W L, Schuetz M, Brady S M, Douglas C J. 2014. BEL1-LIKE HOMEODOMAIN6 and KNOTTED ARABIDOPSIS THALIANA7 interact and regulate secondary cell wall formation via repression of REVOLUTA. The Plant Cell, 26(12):4843-61. doi: 10.1105/tpc.114.128322 URL
[24]	Livak K J, Schmittgen T D. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2^-ΔΔCT method. Methods, 25(4):402-408. pmid: 11846609
[25]	Ma Q, Wang N H, Hao P B, Sun H R, Wang C C, Ma L, Wang H T, Zhang X L, Wei H L, Yu S X. 2019. Genome-wide identification and characterization of TALE superfamily genes in cotton reveals their functions in regulating secondary cell wall biosynthesis. BMC Plant Biology, 19:432. doi: 10.1186/s12870-019-2026-1 URL
[26]	Meister R J, Williams L A, Monfared M M, Gallagher T L, Kraft E A, Nelson C G, Gasser C S. 2004. Definition and interactions of a positive regulatory element of the Arabidopsis INNER NO OUTER promoter. Plant Journal, 37(3):426-438. pmid: 14731261
[27]	Mukherjee K, Brocchieri L, Burglin T R. 2009. A comprehensive classification and evolutionary analysis of plant homeobox genes. Molecular Biology and Evolution, 26(12):2775. doi: 10.1093/molbev/msp201 pmid: 19734295
[28]	Nardmann J, Werr W. 2006. The shoot stem cell niche in Angiosperms:expression patterns of WUS orthologues in rice and maize imply major modifications in the course of mono- and dicot evolution. Molecular Biology and Evolution, 23(12):2492-2504. doi: 10.1093/molbev/msl125 URL
[29]	Pagnussat G C, Yu H J, Sundaresan V. 2007. Cell-fate switch of synergid to egg cell in Arabidopsis eostre mutant embryo sacs arises from misexpression of the BEL1-like homeodomain gene BLH1. The Plant Cell, 19(11):3578-92. doi: 10.1105/tpc.107.054890 URL
[30]	Poulios S, Vlachonasios K E. 2018. Synergistic action of GCN5 and CLAVATA1 in the regulation of gynoecium development in Arabidopsis thaliana. New Phytologist, 220(2):593-608. doi: 10.1111/nph.15303 URL
[31]	Reiser L, Modrusan Z, Margossian L, Samach A, Ohad N, Haughn G W, Fischer R L. 1995. The BELL1 gene encodes a homeodomain protein involved in pattern formation in the Arabidopsis ovule primordium. Cell, 83(5):735-742. pmid: 8521490
[32]	Schoof H, Lenhard M, Haecker A, Mayer K F, Jürgens G, Laux T. 2000. The stem cell population of Arabidopsis shoot meristems in maintained by a regulatory loop between the CLAVATA and WUSCHEL genes. Cell, 100(6):635-644. pmid: 10761929
[33]	Scofield S, Murray J A H. 2006. KNOX gene function in plant stem cell niches. Plant Molecular Biology, 60(6):929-946. doi: 10.1007/s11103-005-4478-y URL
[34]	Skinner D J, Gasser C S. 2009. Expression-based discovery of candidate ovule development regulators through transcriptional profiling of ovule mutants. BMC Plant Biology, 9:29. doi: 10.1186/1471-2229-9-29 pmid: 19291320
[35]	Sloan J, Jana P H, Michael G, Olga E, Andrea G, Gunter S, Klemens W, Irmgard S, Jan U L. 2020. Structural basis for the complex DNA binding behavior of the plant stem cell regulator WUSCHEL. Nature Communications,doi: 10.1038/s41467-020-16024-y. doi: 10.1038/s41467-020-16024-y
[36]	Somssich M, Je B I, Simon R, Jackson D. 2016. CLAVATA-WUSCHEL signaling in the shoot meristem. Development, 143(18):3238-3248. doi: 10.1242/dev.133645 pmid: 27624829
[37]	Song Wei-na, Zhao Sen-sen, Wu Ming-zhu, Wang Gen-fa, Luo Zhao-peng, Wang Chen, Li Ze-feng, Yang Jun, Wang Zhong. 2020. Identification of tobacco BELL gene family and their expression patterns in greenhouses. Tobacco Science & Technology, 53(1):1-11. (in Chinese)
	宋卫娜, 赵森森, 武明珠, 王根发, 罗朝鹏, 王晨, 李泽锋, 杨军, 王中. 2020. 烟草BELL基因家族鉴定及其温室条件下的表达模式分析. 烟草科技, 53(1):1-11.
[38]	Sun B, Ito T. 2015. Regulation of floral stem cell termination in Arabidopsis. Frontiers in Plant Science, 6:17. doi: 10.3389/fpls.2015.00017 pmid: 25699061
[39]	Su Y H, Liu Y B, Bai B, Zhang X S. 2015. Establishment of embryonic shoot-root axis is involved in auxin and cytokinin response during Arabidopsis somatic embryogenesis. Front in Plant Science, 5:792.
[40]	Tan Wen-bo, Li Yu-hua, Xu Qi-jiang. 2008. Regulatory mechanism of stem cell in plant shoot apical meristem. Plant physiology Journal, 44(4):811-816. (in Chinese)
	谭文勃, 李玉花, 徐启江. 2008. 植物茎端分生组织中的茎干细胞调控机制. 植物生理学报, 44(4):811-816.
[41]	Tatyana R, Dolf W. 2014. A roadmap to embryo identity in plants. Trends in Plant Science, 19(11):709-716. doi: 10.1016/j.tplants.2014.06.009 pmid: 25017700
[42]	van Der G E, Laux T, Rensing S A. 2009. The WUS homeobox-containing(WOX)protein family. Genome Biology, 10(12):248. doi: 10.1186/gb-2009-10-12-248 URL
[43]	Wang Y Y, Zhao Y J, Yan M, Zhao H L, Zhang X H, Yuan Z H. 2020. Genome-wide identification and expression analysis of TALE gene family in pomegranate(Punica granatum L.). Agronomy,doi: 10.3390/agronomy10060829. doi: 10.3390/agronomy10060829
[44]	Wang Chu-kun, Han Peng-liang, Wang Yong-mei, Wang Peng-fei, Hu Da-gang. 2019. Genome-wide identification and analysis of apple WUSCHEL-related homeobox(WOX)family genes. Acta Horticulturae Sinica, 46(6) 10:1021-1032.
	王楚堃, 韩朋良, 王咏梅, 王鹏飞, 胡大刚. 2019. 苹果WUSCHEL-related homeobox(WOX)家族基因的鉴定与分析. 园艺学报, 46(6):1021-1032.
[45]	Wetzstein H Y, Ravid N, Wilkins E, Adriana P M. 2011. A morphological and histological characterization of bisexual and male flower types in pomegranate. Journal of the American Society for Horticultural Science, 136(2):83-92. doi: 10.21273/JASHS.136.2.83 URL
[46]	Xu Yun-yuan, Zhong Kang. 2005. Progress in research on plant stem cell organizer gene WUSCHEL. Journal of Plant Physiology and Molecular Biology, 31(5):461-468. (in Chinese)
	徐云远, 种康. 2005. 植物干细胞决定基因WUS的研究进展. 植物生理与分子生物学学报, 31(5):461-468.
[47]	Xu Zhi-hong, Zhong Kang. 2015. Plant cell differentiation and organogenesis. Biejing: Science Press: 246-262. (in Chinese)
	许智宏, 种康. 2015. 植物细胞分化与器官发生. 北京: 科学出版社: 246-262.
[48]	Yamada T, Sasaki Y, Hashimoto K, Nakajima K, Gasser C S. 2016. CORONA,PHABULOSA and PHAVOLUTA collaborate with BELL1 to confine WUSCHEL expression to the nucellus in Arabidopsis ovules. Development, 143(3):422-426.
[49]	Yuan Z H, Fang Y M, Zhang T K, Fei Z J, Han F M, Liu C Y, Liu M, Xiao W, Zhang W J, Wu S, Zhang M W, Ju Y H, Xu H L, Dai H, Liu Y J, Chen Y H, Wang L L, Zhou J Q, Guan D, Yan M, Xia Y H, Huang X B, Liu D Y, Wei H M, Zheng H K. 2018. The pomegranate (Punica granatum L.)genome provides insights into fruit quality and ovule developmental biology. Plant Biotechnology Journal, 16:1363-1374. doi: 10.1111/pbi.2018.16.issue-7 URL
[50]	Yuan Zhao-he.2015. Science and practice of fruit trees in China · pomegranate. Xi’an: Shananxi Science and Technology Press. (in Chinese)
	苑兆和. 2015. 中国果树科学与实践 · 石榴. 西安: 陕西科学技术出版社.
[51]	Zhang Dong-min, Tian Qi-lin, Yang Man-man, Lin Yu-ling, Lai Zhong-xiong. 2015. Cloning of DIWUS from embryogenic callus and its expression analysis during somatic embryogenesis in Dimocarpus longan Lour. Acta Botanica Boreali-Occidentalia Sinica, 35(5):890-897. (in Chinese)
	张冬敏, 田奇琳, 杨曼曼, 林玉玲, 赖钟雄. 2015. 龙眼体细胞胚胎发生过程中D1WUS的克隆与表达分析. 西北植物学报, 35(5):890-897.
[52]	Zhang X, Zong J, Liu J, Yin J, Zhang D. 2010. Genome-wide analysis of WOX gene family in rice,sorghum,maize,Arabidopsis and poplar. Journal of Integrative Plant Biology, 52(11):1016-1026. doi: 10.1111/j.1744-7909.2010.00982.x pmid: 20977659
[53]	Zhang Z J, Tucker E, Hermann M, Laux T. 2017. A molecular framework for the embryonic initiation of shoot meristem stem cells. Developmental Cell, 40(3):264-277. doi: 10.1016/j.devcel.2017.01.002 URL
[54]	Zhao Y J, Liu C Y, Ge D P, Yan M, Ren Y, Huang X B, Yuan Z H. 2020. Genome-wide identification and expression of YABBY genes family during flower development in Punica granatum L. Gene, 752:44784.
[55]	Zhou Yang, Lui Wei, Sun Shi, Han Tian-fu, Hou Wen-sheng. 2014. Molecular cloning and expression of GmWUS related to regeneration in soybean. Chinese Journal of Oil Crop Sciences, 36(6):707-712. (in Chinese)
	周扬, 刘薇, 孙石, 韩天富, 侯文胜. 2014. 大豆再生相关基因GmWUS的克隆及表达分析. 中国油料作物学报, 36(6):707-712.

Cloning and Spatiotemporal Expression Analysis of PgWUS and PgBEL1 in Punica granatum

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