Welcome to Acta Agronomica Sinica，

Acta Agronomica Sinica ›› 2019, Vol. 45 ›› Issue (8): 1146-1157.doi: 10.3724/SP.J.1006.2019.84124

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Identification and function analysis of AVP1, VHA-a2, and VHA-a3 genes in Brassica napus L.

YAO Jun-Yue1,2,HUA Ying-Peng1,2,ZHOU Ting1,2,WANG Tao1,2,SONG Hai-Xing1,2,GUAN Chun-Yun3,ZHANG Zhen-Hua1,2,*()   

  1. 1 College of Resource and Environment, Hunan Agricultural University, Changsha 410128, Hunan, China
    2 Southern Regional Collaborative Innovation Center for Grain and Oil Crops in China, Changsha 410128, Hunan, China
    3 Hunan Branch, National Oil Crops Improvement Center, Changsha 410128, Hunan, China
  • Received:2018-09-20 Accepted:2019-04-15 Online:2019-08-12 Published:2019-07-16
  • Contact: Zhen-Hua ZHANG E-mail:zhzh1468@163.com
  • Supported by:
    This study was supported by the National Key R&D Program of China(2018YFD0200901);This study was supported by the National Key R&D Program of China(2018YFD0200906);the China Agriculture Research System(CARS-13)

RichHTML354

16

PDF

552

Abstract:

Vacuoles play an important role in regulating plant cell differentiation and growth. AVP1, VHA-a2, and VHA-a3 genes are important carrier elements for regulating the acid-base balance inside and outside the vacuole of plants and the energy provided by ion transport. In this study, nine BnaAVP1, three BnaVHA-a2, and four BnaVHA-a3 genes were identified in Brassica napus genome database and NCBI plant genome annotation database by using the known AVP1, VHA-a2, and VHA-a3 genes of Arabidopsis thaliana as reference sequences. Bioinformatics studies on copy number variation, molecular characteristics, transmembrane domain, conserved motifs, chromosome localization, phylogenetic tree construction, secondary structure and three- dimensional structure prediction of proteins and high-throughput transcriptome sequencing were carried out, indicating that the number of genes of BnaAVP1 and BnaVHA-a3 was significantly higher than that of B. oleracea and B. rapa; the AVP1, VHA-a2 and VHA-a3 proteins of B. napus belonged to a stable protein composed of acidic amino acids. The analysis of evolutionary selection ability showed that after low nitrogen treatment, BnaAVP1 genes were mainly expressed in shoots, and the expression of BnaAVP1 genes was down-regulated in shoots after three hours of low nitrogen treatment, and up-regulated in roots after 72 hours of low nitrogen treatment. BnaVHA-a2 and BnaVHA-a3 genes were expressed in both shoots and roots, while BnaVHA-a2 genes were up-regulated after 72 hours of low nitrogen treatment, and BnaVHA-a3 genes were down-regulated after three hours of low nitrogen treatment. After low phosphorus treatment, the expression of most BnaAVP1 genes was up-regulated in roots, and no difference in shoots. The expression of BnaVHA-a2 genes was almost no difference, and that of BnaVHA-a3 genes up-regulated in both shoots and roots. This result lays a foundation for further study on the biological functions of AVP1, VHA-a2, and VHA-a3 genes in B. napus and the molecular mechanism that AVP1, VHA-a2, and VHA-a3 proteins hydrolyze ATP to provide energy for plant metabolism, and provides references for other species family genetic bioinformatics studies that are known to have large amounts of data.

Key words: Brassica napus L, AVP gene, VHA gene, bioinformatics

Fig. 1

Copy number variation of AVP1, VHA-a2, and VHA-a3 genes in Brassica napus, B. rapa, and B. olerace The number at the top of the histogram is the number of genes copied for that species."

Table 1

Information of AVP1, VHA-a2, and VHA-a3 genes in Brassica napus"

基因名称
Gene name		 基因库编号
Database No.		 基因编号
Gene ID		 基因长度
CDS (bp)		 氨基酸长度
Amino acids (aa)		 分子量
MW (kD)		 理论等电点
pI		 亲水性指数
GRAVY
BnaAVP1-1 GSBRNA2T00010424001 BnaA09g56670D 2409 802 83.97 5.06 0.620
BnaAVP1-2 GSBRNA2T00033628001 BnaA06g38620D 1821 606 64.61 5.00 0.281
BnaAVP1-3 GSBRNA2T00136659001 BnaC08g38730D 3525 1174 126.46 6.29 0.269
BnaAVP1-4 GSBRNA2T00139755001 BnaC08g16730D 2310 769 80.59 5.03 0.611
BnaAVP1-5 GSBRNA2T00068861001 BnaC06g40530D 2310 769 80.61 5.00 0.574
BnaAVP1-6 GSBRNA2T00054245001 BnaC05g11930D 2316 771 80.83 5.08 0.618
BnaAVP1-7 GSBRNA2T00103071001 BnaA08g23780D 2310 769 80.52 5.03 0.616
BnaAVP1-8 GSBRNA2T00146278001 BnaA07g35600D 2310 769 80.70 5.01 0.572
BnaAVP1-9 GSBRNA2T00062272001 BnaA06g10340D 2316 771 80.83 5.08 0.621
BnaVHA-a2-1 GSBRNA2T00137010001 BnaC08g35720D 2466 821 92.97 5.61 0.060
BnaVHA-a2-2 GSBRNA2T00108093001 BnaC04g33510D 2472 823 92.96 5.61 0.056
BnaVHA-a2-3 GSBRNA2T00090255001 BnaA04g12160D 2460 819 92.80 5.56 0.059
BnaVHA-a3-1 GSBRNA2T00061106001 BnaC03g60340D 2463 820 92.85 5.86 0.041
BnaVHA-a3-2 GSBRNA2T00075109001 BnaC01g00150D 2457 818 92.50 5.80 0.080
BnaVHA-a3-3 GSBRNA2T00150888001 BnaA08g16770D 2460 819 92.79 5.75 0.043
BnaVHA-a3-4 GSBRNA2T00073949001 BnaA01g05300D 2454 817 92.32 5.66 0.071

Fig. 2

Synonymous nucleotide substitution rates (Ks) and non-synonymous nucleotide substitution rates (Ka) of the BnaAVP1, BnaVHA-a2, and BnaVHA-a3 proteins in Brassica napus L. "

Fig. 3

Chromosome localization of the BnaAVP1s, BnaVHA-a2s, and BnaVHA-a3s of Brassica napus"

Fig. 4

Distribution of conserved motifs in BnaAVP1s, BnaVHA-a2s, and BnaVHA-a3s of Brassica napus"

Fig. 5

Exon-intron structure of BnaAVP1, BnaVHA-a2, and BnaVHA-a3 of Brassica napus"

Fig. 6

Evolutionary relationships of BnaAVP1, BnaVHA-a2, and BnaVHA-a3 proteins of Brassica napus Blue represents the BnaAVP1 family, green represents the BnaVHA-a3 family, and red represents the BnaVHA-a2 family."

Fig. 7

Secondary structure analysis of BnaVHA-a2-1 protein GOR4 (A) and PSIPRED (B) are two tools for the secondary structure analysis."

Fig. 8

Prediction of three-dimensional structure of BnaAVP1, BnaVHA-a2, and BnaVHA-a3 protein A: BnaVHA-a2-1; B: BnaVHA-a3-1; C: BnaAVP1-1."

Fig. 9

Transcriptome analysis of BnaAVP1, BnaVHA-a2, and BnaVHA-a3 family genes in Brassica napus S: shoot; R: root; 0, 3, and 72 (h) represent the time course of nitrogen deficiency. The results represent the average of three biological replicates. * represent significant differences. (A): BnaVHA-a2s; (B): BnaVHA-a3s; (C): BnaAVP1s."

Fig. 10

Transcriptome analysis of BnaAVP1, BnaVHA-a2, and BnaVHA-a3 family genes in Brassica napus S: shoot; R: root; CK represent normal condition and LP represent low phosphorus stress. The results represent the average of three biological replicates. * represent significant differences. (A): BnaVHA-a2s; (B): BnaVHA-a3s; (C): BnaAVP1s."

[1] Sze H, Schumacher K, Müller M L, Padmanaban S, Taiz L . A simple nomenclature for a complex proton pump: VHA genes encode the vacuolar H +-ATPase. Trends Plant Sci, 2002,7:157-161.
[2] Sze H, Li X, Palmgren M G . Energization of plant cell membranes by H+-pumping ATPases: regulation and biosynthesis. Plant Cell, 1999,11:677-689.
[3] Schumacher K . pH in the plant endomembrane system: an import and export business. Curr Opin Plant Biol, 2014,22:71-76.
doi: 10.1016/j.pbi.2014.09.005
[4] Drozdowicz Y M, Rea P A . Vacuolar H+ pyrophosphatases: from the evolutionary backwaters into the mainstream. Trends Plant Sci, 2001,6:206-211.
doi: 10.1016/S1360-1385(01)01923-9
[5] Maeshima M . Tonoplast transporters: organization and function. Annu Rev Plant Biol, 2001,52:469-497.
doi: 10.1146/annurev.arplant.52.1.469
[6] Bayer P E, Hurgobin B, Golicz A A, Chan C K K, Yuan Y, Lee H, Zou J . Assembly and comparison of two closely related Brassica napus genomes. Plant Biotechnol J, 2017,15:1602-1610.
doi: 10.1111/pbi.2017.15.issue-12
[7] Altschul S F, Gish W, Miller W, Myers E W, Lipman D J . Basic local alignment search tool. J Mol Biol, 1990,215:403-410.
doi: 10.1016/S0022-2836(05)80360-2
[8] Finn R D, Bateman A, Clements J, Coggill P, Eberhardt R Y, Eddy S R, Sonnhammer E L . Pfam: the protein families database. Nucl Acids Res, 2013,42:D222-D230.
[9] Letunic I, Doerks T, Bork P . SMART: recent updates, new developments and status in 2015. Nucl Acids Res, 2014,43:D257-D260.
[10] Gasteiger E, Gattiker A, Hoogland C, Ivanyi I, Appel R D, Bairoch A . ExPASy: the proteomics server for in-depth protein knowledge and analysis. Nucl Acids Res, 2003,31:3784-3788.
doi: 10.1093/nar/gkg563
[11] Bailey T, Lelka N . Fitting a mixture model by expectation maximization to discover motifs in biopolymers. Proc Int Conf Intell Syst Mol Biol, 1994,4:28-36.
[12] Aiyar A . The use of CLUSTAL W and CLUSTAL X for multiple sequence alignment. Methods Mol Biol, 2000,132:221-241.
[13] Kumar S, Nei M, Dudley J, Tamura K . MEGA: a biologist-centric software for evolutionary analysis of DNA and protein sequences. Brief Bioinform, 2008,9:299-306.
doi: 10.1093/bib/bbn017
[14] Krzywinski M I, Schein J E, Birol I, Connors J, Gascoyne R, Horsman D, Marra M A . Circos: an information aesthetic for comparative genomics. Genome Res, 2009,19:1639-1645.
doi: 10.1101/gr.092759.109
[15] Combet C, Blanchet C, Geourjon C, Deleage G . NPS@: network protein sequence analysis. Trends Biochem Sci, 2000,25:147-150.
doi: 10.1016/S0968-0004(99)01540-6
[16] Jones D T . Protein secondary structure prediction based on position-specific scoring matrices. J Mol Biol, 1999,292:195-202.
doi: 10.1006/jmbi.1999.3091
[17] Buchan D W, Minneci F, Nugent T C, Bryson K, Jones D T . Scalable web services for the PSIPRED Protein Analysis Workbench. Nucl Acids Res, 2013,41:W349-W357.
doi: 10.1093/nar/gkt381
[18] Mezulis S, Sternberg M J, Kelley L A . PhyreStorm: A web server for fast structural searches against the PDB. J Mol Biol, 2016,428:702-708.
doi: 10.1016/j.jmb.2015.10.017
[19] Hoagland D R, Arnon D I . The water culture method for growing plants without soil. Calif Agric Exp Stn Circ, 1950,347:4-32.
[20] Morin R D, Bainbridge M, Fejes A, Hirst M, Krzywinski M, Pugh T J, Marra M A . Profiling the HeLa S3 transcriptome using randomly primed cDNA and massively parallel short-read sequencing. Biotechniques, 2008,45:81-94.
doi: 10.2144/000112900
[21] Hua Y, Feng Y, Zhou T, Xu F . Genome-scale mRNA transcriptomic insights into the responses of oilseed rape (Brassica napus L.) to varying boron availabilities. Plant Soil, 2017,416:205-225.
[22] Eisen M B, Spellman P T, Brown P O, Botstein D . Cluster analysis and display of genome-wide expression patterns. Proc Natl Acad Sci USA, 1998,95:14863-14868.
doi: 10.1073/pnas.95.25.14863
[23] Kohl M, Wiese S, Warscheid B . Cytoscape: software for visualization and analysis of biological networks. Methods Mol Biol, 2011,696:291-303.
doi: 10.1007/978-1-60761-987-1
[24] Nekrutenko A, Makova K D, Li W H . The KA/KS ratio test for assessing the protein-coding potential of genomic regions: an empirical and simulation study. Genome Res, 2002,12:198-202.
[25] Segami S, Nakanishi Y, Sato M H, Maeshima M . Quantification, organ-specific accumulation and intracellular localization of type II H+-pyrophosphatase in Arabidopsis thaliana. Plant Cell Physiol, 2010,51:1350-1360.
doi: 10.1093/pcp/pcq096
[26] Li J, Yang H, Peer W A, Richter G, Blakeslee J, Bandyopadhyay A, Krizek B . Arabidopsis H+-PPase AVP1 regulates auxin- mediated organ development . Science, 2005,310:121-125.
doi: 10.1126/science.1115711
[27] Gaxiola R A, Palmgren M G, Schumacher K . Plant proton pumps. Febs Lett, 2007,581:2204-2214.
doi: 10.1016/j.febslet.2007.03.050
[28] Beyenbach K W, Wieczorek H . The V-type H+ ATPase: molecular structure and function, physiological roles and regulation . J Exp Biol, 2006,209:577-589.
doi: 10.1242/jeb.02014
[29] Narasimhan M L, Binzel M L, Perez-Prat E, Chen Z, Nelson D E, Singh N K, Hasegawa P M . NaCl regulation of tonoplast ATPase 70-kilodalton subunit mRNA in tobacco cells. Plant Physiol, 1991,97:562-568.
doi: 10.1104/pp.97.2.562
[30] Binzel M L, Dunlap J R . Abscisic acid does not mediate NaCl-induced accumulation of 70-kDa subunit tonoplast H+-ATPase message in tomato . Planta, 1995,197:563-568.
[31] Yamashita K, Kasai M, Ezaki B, Shibasaka M, Yamamoto Y, Matsumoto H, Sasakawa H . Stimulation of H+ extrusion and plasma membrane H+-ATPase activity of barley roots by ammonium-treatment . Soil Sci Plant Nutr, 1995,41:133-140.
doi: 10.1080/00380768.1995.10419566
[32] 周金泉, 张明超, 魏志军, 胡军, 朱毅勇 . 高粱分泌硝化抑制物对羟基苯丙酸与质子泵的关系研究. 土壤学报, 2015,52:620-627.
Zhou J Q, Zhang M C, Wei Z J, Hu J, Zhu Y Y . Relationship between exudation of nitrification inhibitor MHPP and plasma membrane proton pump of sorghum root. J Soil, 2015,52:620-627 (in Chinese with English abstract).
[33] De Angeli A, Monachello D, Ephritikhine G, Frachisse J M, Thomine S, Gambale F, Barbier-Brygoo H . The nitrate/proton antiporter AtCLCa mediates nitrate accumulation in plant vacuoles. Nature, 2006,442:939-942.
[34] 黄海涛, 张振华, 宋海星, 刘强, 荣湘民, 彭建伟, 官春云 . 基于液泡膜质子泵的硝态氮再利用研究进展. 生态学杂志, 2012,31:731-737.
Huang H T, Zhang Z H, Song H X, Liu Q, Rong X M, Peng J W, Guan C Y . Nitrate nitrogen reutilization based on proton pump of vacuole membrane: a review. J Ecol, 2012,31:731-737 (in Chinese with English abstract).
[1] WU Yan-Fei, HU Qin, ZHOU Qi, DU Xue-Zhu, SHENG Feng. Genome-wide identification and expression analysis of Elongator complex family genes in response to abiotic stresses in rice [J]. Acta Agronomica Sinica, 2022, 48(3): 644-655.
[2] ZHAO Gai-Hui, LI Shu-Yu, ZHAN Jie-Peng, LI Yan-Bin, SHI Jia-Qin, WANG Xin-Fa, WANG Han-Zhong. Mapping and candidate gene analysis of silique number mutant in Brassica napus L. [J]. Acta Agronomica Sinica, 2022, 48(1): 27-39.
[3] JIAN Hong-Ju, SHANG Li-Na, JIN Zhong-Hui, DING Yi, LI Yan, WANG Ji-Chun, HU Bai-Geng, Vadim Khassanov, LYU Dian-Qiu. Genome-wide identification and characterization of PIF genes and their response to high temperature stress in potato [J]. Acta Agronomica Sinica, 2022, 48(1): 86-98.
[4] HUANG Ning, HUI Qian-Long, FANG Zhen-Ming, LI Shan-Shan, LING Hui, QUE You-Xiong, YUAN Zhao-Nian. Identification, localization and expression analysis of beta-carotene isomerase gene family in sugarcane [J]. Acta Agronomica Sinica, 2021, 47(5): 882-893.
[5] ZHANG Chun, ZHAO Xiao-Zhen, PANG Cheng-Ke, PENG Men-Lu, WANG Xiao-Dong, CHEN Feng, ZHANG Wei, CHEN Song, PENG Qi, YI Bin, SUN Cheng-Ming, ZHANG Jie-Fu, FU Ting-Dong. Genome-wide association study of 1000-seed weight in rapeseed (Brassica napus L.) [J]. Acta Agronomica Sinica, 2021, 47(4): 650-659.
[6] XIE Pan, LIU Wei, KANG Yu, HUA Wei, QIAN Lun-Wen, GUAN Chun-Yun, HE Xin. Identification and relative expression analysis of CBF gene family in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(12): 2394-2406.
[7] LI Peng, LIU Che, SONG Hao, YAO Pan-Pan, SU Pei-Lin, WEI Yao-Wei, YANG Yong-Xia, LI Qing-Chang. Identification and analysis of non-specific lipid transfer protein family in tobacco [J]. Acta Agronomica Sinica, 2021, 47(11): 2184-2198.
[8] HUANG Xiao-Fang,BI Chu-Yun,SHI Yuan-Yuan,HU Yun-Zhuo,ZHOU Li-Xiang,LIANG Cai-Xiao,HUANG Bi-Fang,XU Ming,LIN Shi-Qiang,CHEN Xuan-Yang. Discovery and analysis of NBS-LRR gene family in sweet potato genome [J]. Acta Agronomica Sinica, 2020, 46(8): 1195-1207.
[9] FENG Tao,TAN Hui,GUAN Mei,GUAN Chun-Yun. Mechanism of BnaBZR1 and BnaPIF4 regulating photosynthetic efficiency in oilseed rape (Brassica napus L.) under poor light [J]. Acta Agronomica Sinica, 2020, 46(8): 1146-1156.
[10] ZHENG Qing-Lei,YU Chen-Jing,YAO Kun-Cun,HUANG Ning,QUE You-Xiong,LING Hui,XU Li-Ping. Cloning and expression analysis of sugarcane Fe/S precursor protein gene ScPetC [J]. Acta Agronomica Sinica, 2020, 46(6): 844-857.
[11] SUN Cheng-Ming,CHEN Feng,CHEN Song,PENG Qi,ZHANG Wei,YI Bin,ZHANG Jie-Fu,FU Ting-Dong. Genome-wide association study of seed number per silique in rapeseed (Brassica napus L.) [J]. Acta Agronomica Sinica, 2020, 46(01): 147-153.
[12] SUN Cheng-Ming,CHEN Song,PENG Qi,ZHANG Wei,YI Bin,ZHANG Jie-Fu,FU Ting-Dong. Genome-wide association study of silique length in rapeseed (Brassica napus L.) [J]. Acta Agronomica Sinica, 2019, 45(9): 1303-1310.
[13] SUN Ting-Ting,WANG Wen-Ju,LOU Wen-Yue,LIU Feng,ZHANG Xu,WANG Ling,CHEN Yu-Feng,QUE You-Xiong,XU Li-Ping,LI Da-Mei,SU Ya-Chun. Cloning and expression analysis of sugarcane lipoxygenase gene ScLOX1 [J]. Acta Agronomica Sinica, 2019, 45(7): 1002-1016.
[14] Jing LI,Jin-Yao YAN,Wen-Shi HU,Xiao-Kun LI,Ri-Huan CONG,Tao REN,Jian-Wei LU. Effects of combined application of nitrogen and potassium on seed yield and nitrogen utilization of winter oilseed rape (Brassica napus L.) [J]. Acta Agronomica Sinica, 2019, 45(6): 941-948.
[15] Gui-Hong LIANG,Ying-Peng HUA,Ting ZHOU,Qiong LIAO,Hai-Xing SONG,Zhen-Hua ZHANG. Bioinformatics analysis and response to nitrate-cadmium stress of NRT1.5 and NRT1.8 family genes in Brassica napus [J]. Acta Agronomica Sinica, 2019, 45(3): 365-380.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
Add: No. 80 Xueyuan South Rd, Beijing 100081, P. R. China E-mail: zwxb301@caas.cn
Supported by Beijing Magtech Co.Ltd