Genome-wide Identification of U-box E3 Ubiquitin Ligase Gene Family in Setaria italica and Response Analysis to Abiotic Stress

doi:10.13560/j.cnki.biotech.bull.1985.2024-0983

Abstract

Abstract:

Objective The ubiquitin ligase U-box (plant U-box protein, PUB) gene family (SiPUB) of Setaria italica was systematically identified based on the whole genome level, which may provide theoretical support for the study of the biological function of the ubiquitin ligase U-box gene family of S. italica. Method The U-box gene family members of S. italica were identified by the whole genome of the U-box conserved Pfam sequence. The phylogenetic tree construction, subcellular localization prediction, physical and chemical properties such as relative molecular mass and isoelectric point of the predicted protein, and Scaffold localization map of the family were analyzed. The expressions of U-box gene family members in S. italica under different stress treatments were studied by real-time fluorescence quantitative PCR (RT-qPCR). Result There were 71 PUB gene family members in the genome of S. italica, which were unevenly distributed on 8 chromosomes. Phylogenetic analysis showed that SiPUB genes were distributed in subfamilies I to Ⅸ. Promoter cis-acting element analysis showed that U-box genes were related to plant growth and development, stress and hormone regulation. The results of fluorescence quantitative PCR showed that SiPUBs were differentially expressed in various parts of S. italica. The expression patterns of different domains of S. italicaU-box gene family under stress were analyzed. The expressions of genes containing U-box domain changed significantly under low temperature and drought, and the expressions of genes containing U-box+HEAT_2 domain did not change significantly under three stresses. The expressions of genes containing U-box+Pkinase and other domains increased and decreased under different stresses, indicating that each domain played different roles in plant environmental response and gene regulation. Plants responded to abiotic stress by regulating the expression of SiPUBs. Conclusion A total of 71 SiPUBs genes are identified, and the expression patterns of different subfamilies under abiotic stress are different, indicating that SiPUBs play an important regulatory role in the response of S. italica to abiotic stress. The results of this study can provide reference for the analysis of stress resistance function of SiPUBs.

Key words: Setaria italica, E3 ubiquitin ligase, U-box gene family, low temperature stress, drought stress, MeJA stress

HE Wei, LI Jun-yi, LI Xin-ni, MA Xue-hua, XING Yuan, CAO Xiao-ning, QIAO Zhi-ju, LIU Si-chen. Genome-wide Identification of U-box E3 Ubiquitin Ligase Gene Family in Setaria italica and Response Analysis to Abiotic Stress[J]. Biotechnology Bulletin, 2025, 41(5): 104-118.

Figures/Tables 12

References 38

1	刁现民. 育种创新造就谷子种业新发展 [J]. 中国种业, 2022(4): 4-7.
	Diao XM. Breeding innovation creates new development of millet seed industry [J]. China Seed Ind, 2022(4): 4-7.
2	邢媛, 宋健, 李俊怡, 等. 谷子AP基因家族鉴定及其对非生物胁迫的响应分析 [J]. 生物技术通报, 2023, 39(11): 238-251.
	Xing Y, Song J, Li JY, et al. Identification of AP gene family and its response analysis to abiotic stress in Setaria italica [J]. Biotechnol Bull, 2023, 39(11): 238-251.
3	王建雄. 山西吕梁地区近50年气候变化与主要气象灾害的研究 [D]. 兰州: 兰州大学, 2012.
	Wang JX. Study on climate change and main meteorological disasters in Luliang area of Shanxi province in recent 50 years [D]. Lanzhou: Lanzhou University, 2012.
4	Craig A, Ewan R, Mesmar J, et al. E3 ubiquitin ligases and plant innate immunity [J]. J Exp Bot, 2009, 60(4): 1123-1132.
5	Marshall RS, Vierstra RD. Dynamic regulation of the 26S proteasome: from synthesis to degradation [J]. Front Mol Biosci, 2019, 6: 40.
6	Vierstra RD. The ubiquitin-26S proteasome system at the nexus of plant biology [J]. Nat Rev Mol Cell Biol, 2009, 10(6): 385-397.
7	Patterson C. A new Gun in town: the U box is a ubiquitin ligase domain [J]. Sci STKE, 2002, 2002(116): pe4.
8	Koegl M, Hoppe T, Schlenker S, et al. A novel ubiquitination factor, E4, is involved in multiubiquitin chain assembly [J]. Cell, 1999, 96(5): 635-644.
9	Wiborg J, O'Shea C, Skriver K. Biochemical function of typical and variant Arabidopsis thaliana U-box E3 ubiquitin-protein ligases [J]. Biochem J, 2008, 413(3): 447-457.
10	曹英豪. 水稻U-box基因家族的特征及转录表达模式分析 [D]. 保定: 河北农业大学, 2010.
	Cao YH. Characteristics and transcriptional expression patterns of U-box gene family in rice [D]. Baoding: Hebei Agricultural University, 2010.
11	郑兴卫, 邵麟惠, 李聪. 蒺藜苜蓿全基因组中U-box基因家族的筛选与特征分析 [J]. 草业学报, 2015, 24(8): 130-141.
	Zheng XW, Shao LH, Li C. Genome-wide screening and characterization of the U-box gene family in Medicago truncatula [J]. Acta Prataculturae Sin, 2015, 24(8): 130-141.
12	李菲, 何小红, 龚记熠, 等. 番茄基因组中U-box基因家族的鉴定与分析 [J]. 分子植物育种, 2018, 16(11): 3468-3476.
	Li F, He XH, Gong JY, et al. Identification and analysis of U-box gene family in tomato genome [J]. Mol Plant Breed, 2018, 16(11): 3468-3476.
13	Wang N, Liu YP, Cong YH, et al. Genome-wide identification of soybean U-box E3 ubiquitin ligases and roles of GmPUB8 in negative regulation of drought stress response in Arabidopsis [J]. Plant Cell Physiol, 2016, 57(6): 1189-1209.
14	Yu YH, Li XZ, Guo DL, et al. Genome-wide identification and analysis of the U-box family of E3 ligases in grapevine [J]. Russ J Plant Physiol, 2016, 63(6): 835-848.
15	Cho SK, Ryu MY, Song C, et al. Arabidopsis PUB22 and PUB23 are homologous U-Box E3 ubiquitin ligases that play combinatory roles in response to drought stress [J]. Plant Cell, 2008, 20(7): 1899-1914.
16	Cho SK, Chung HS, Ryu MY, et al. Heterologous expression and molecular and cellular characterization of CaPUB1 encoding a hot pepper U-Box E3 ubiquitin ligase homolog [J]. Plant Physiol, 2006, 142(4): 1664-1682.
17	Qin Q, Wang YX, Huang LY, et al. A U-box E3 ubiquitin ligase OsPUB67 is positively involved in drought tolerance in rice [J]. Plant Mol Biol, 2020, 102(1/2): 89-107.
18	Li W, Ahn IP, Ning YS, et al. The U-Box/ARM E3 ligase PUB13 regulates cell death, defense, and flowering time in Arabidopsis [J]. Plant Physiol, 2012, 159(1): 239-250.
19	Marquis V, Smirnova E, Graindorge S, et al. Broad-spectrum stress tolerance conferred by suppressing jasmonate signaling attenuation in Arabidopsis jasmonic acid oxidase mutants [J]. Plant J, 2022, 109(4): 856-872.
20	马李广, 张贺龙, 庞小可, 等. 白菜bZIP转录因子基因家族应答春化反应关键基因表达分析 [J]. 江苏农业学报, 2022, 38(3): 765-774.
	Ma LG, Zhang HL, Pang XK, et al. Genome-wide identification of bZIP transcription factor gene family in Brassica rapa and its association with vernalization [J]. Jiangsu J Agric Sci, 2022, 38(3): 765-774.
21	桂尚枝, 尹倩, 温宏伟, 等. 乌菜春化相关BcGRAS基因鉴定及表达分析 [J]. 植物生理学报, 2022, 58(12): 2273-2285.
	Gui SZ, Yin Q, Wen HW, et al. Genome-wide identification and expression analysis of BcGRAS genes responded to vernalization in Wucai (Brassica campestris ssp. chinensis var. rosularis) [J]. Plant Physiol J, 2022, 58(12): 2273-2285.
22	陈国户, 庞小可, 李广, 等. 白菜NAC基因家族全基因组鉴定及其应答春化反应的表达分析 [J]. 南京农业大学学报, 2022, 45(4): 656-665.
	Chen GH, Pang XK, Li G, et al. Genome-wide identification of NAC gene family in Brassica rapa and its expression analysis of response to vernalization [J]. J Nanjing Agric Univ, 2022, 45(4): 656-665.
23	Farmer LM, Book AJ, Lee KH, et al. The RAD23 family provides an essential connection between the 26S proteasome and ubiquitylated proteins in Arabidopsis [J]. Plant Cell, 2010, 22(1): 124-142.
24	Yee D, Goring DR. The diversity of plant U-box E3 ubiquitin ligases: from upstream activators to downstream target substrates [J]. J Exp Bot, 2009, 60(4): 1109-1121.
25	Zhang JZ. Evolution by gene duplication: an update [J]. Trends Ecol Evol, 2003, 18(6): 292-298.
26	Cannon SB, Mitra A, Baumgarten A, et al. The roles of segmental and tandem gene duplication in the evolution of large gene families in Arabidopsis thaliana [J]. BMC Plant Biol, 2004, 4: 10.
27	Liu JL, Li W, Ning YS, et al. The U-Box E3 ligase SPL11/PUB13 is a convergence point of defense and flowering signaling in plants [J]. Plant Physiol, 2012, 160(1): 28-37.
28	李秋月, 张亚飞, 彭洁, 等. 柑橘U-box基因家族的鉴定及表达分析 [J]. 中国农业科学, 2019, 52(11): 1942-1960.
	Li QY, Zhang YF, Peng J, et al. Genome wide identification and expression analysis of the U-box gene family in Citrus [J]. Sci Agric Sin, 2019, 52(11): 1942-1960.
29	陈曙, 张彧, 陈卓, 等. 玉米泛素连接酶U-box基因家族的全基因组鉴定及表达分析 [J]. 西南农业学报, 2022, 35(3): 481-490.
	Chen S, Zhang Y, Chen Z, et al. Genome-wide identification and expression analysis of ubiquitin ligase U-box gene family in maize [J]. Southwest China J Agric Sci, 2022, 35(3): 481-490.
30	齐晨辉, 赵先炎, 韩朋良, 等. 苹果U-box型E3泛素连接酶MdPUB24的耐盐性和ABA敏感性鉴定 [J]. 园艺学报, 2017, 44(12): 2255-2264.
	Qi CH, Zhao XY, Han PL, et al. Functional identification of salt tolerance and ABA sensitivity of apple U-box E3 ubiquitin ligase MdPUB24 [J]. Acta Hortic Sin, 2017, 44(12): 2255-2264.
31	Liu YC, Wu YR, Huang XH, et al. AtPUB19, a U-box E3 ubiquitin ligase, negatively regulates abscisic acid and drought responses in Arabidopsis thaliana [J]. Mol Plant, 2011, 4(6): 938-946.
32	Tang X, Ghimire S, Liu WG, et al. Potato E3 ubiquitin ligase PUB27 negatively regulates drought tolerance by mediating stomatal movement [J]. Plant Physiol Biochem, 2020, 154: 557-563.
33	Howe GA, Yoshida Y. Evolutionary origin of JAZ proteins and jasmonate signaling [J]. Mol Plant, 2019, 12(2): 153-155.
34	Dunn MA, White AJ, Vural S, et al. Identification of promoter elements in a low-temperature-responsive gene (blt4.9) from barley (Hordeum vulgare L.) [J]. Plant Mol Biol, 1998, 38(4): 551-564.
35	Dvir A, Conaway JW, Conaway RC. Mechanism of transcription initiation and promoter escape by RNA polymerase II [J]. Curr Opin Genet Dev, 2001, 11(2): 209-214.
36	Potenza C, Aleman L, Sengupta-Gopalan C. Targeting transgene expression in research, agricultural, and environmental applications: Promoters used in plant transformation [J]. Vitro Cell Dev Biol Plant, 2004, 40(1): 1-22.
37	Singh K, Foley RC, Oñate-Sánchez L. Transcription factors in plant defense and stress responses [J]. Curr Opin Plant Biol, 2002, 5(5): 430-436.
38	丁雪峰, 刘鸿艳, 罗利军. 水稻OsGRAS1启动子的克隆及多样性分析 [J]. 上海农业学报, 2010, 26(4): 8-14.
	Ding XF, Liu HY, Luo LJ. Cloning and diversity analysis of the OsGRAS1 promoter in rice [J]. Acta Agric Shanghai, 2010, 26(4): 8-14.

引物名称 Primer name	基因序列 Gene sequence （5′‒3′）
25S-F	AGGCAACAGAAACTCCATACG
25S-R	ATGGCATAGCATTCATCACG
SiPUB2-F	GGAGTCACTGCTCCGCAAT
SiPUB2-R	CTTCTCGTCAGCGGGGC
SiPUB3-F	ACAGCGGAAAGAGGCTCATC
SiPUB3-R	CGAGCGACATCAAGTTTGGC
SiPUB8-F	CGGGCTTGAAGAGAAGCTGA
SiPUB8-R	ACTCGTGCAGGAGAAAGTCG
SiPUB24-F	GTTACTCCTGAGGTCACTTG
SiPUB24-R	TCCACCTGTAACATTGTCTG
SiPUB29-F	CCATTGGTGGTTCAGACATA
SiPUB29-R	AGGGAGTCCAAATATCCGTA
SiPUB38-F	TCAGATTCATTCTCCCAAGC
SiPUB38-R	CCTGAACTTTCTGAAGCAAC
SiPUB46-F	TCAGAGAAATTGACGAGGAG
SiPUB46-R	ATAATCCTGTCAGCAGGTTG
SiPUB49-F	GGAGCGAGATGATGCGGTTA
SiPUB49-R	AGTGAACGGCTGAAGACCTG
SiPUB55-F	CCAGCATCGAGTCCTGGGTC
SiPUB55-R	ACGCACCACTCCTGGATGA
SiPUB59-F	CAACAAGCCCAAGAGGGACT
SiPUB59-R	GCTGGCTTCCCAGTTACCAT
SiPUB64-F	CTCACAGGAACAAAGCGTGC
SiPUB64-R	CACGCCTCAATGTGCTGTTC
SiPUB70-F	CTGTTCCTGTGCCCCATCTC
SiPUB70-R	CCGTACCCGAAGATCCACTG

引物名称 Primer name	基因序列 Gene sequence （5′‒3′）
25S-F	AGGCAACAGAAACTCCATACG
25S-R	ATGGCATAGCATTCATCACG
SiPUB2-F	GGAGTCACTGCTCCGCAAT
SiPUB2-R	CTTCTCGTCAGCGGGGC
SiPUB3-F	ACAGCGGAAAGAGGCTCATC
SiPUB3-R	CGAGCGACATCAAGTTTGGC
SiPUB8-F	CGGGCTTGAAGAGAAGCTGA
SiPUB8-R	ACTCGTGCAGGAGAAAGTCG
SiPUB24-F	GTTACTCCTGAGGTCACTTG
SiPUB24-R	TCCACCTGTAACATTGTCTG
SiPUB29-F	CCATTGGTGGTTCAGACATA
SiPUB29-R	AGGGAGTCCAAATATCCGTA
SiPUB38-F	TCAGATTCATTCTCCCAAGC
SiPUB38-R	CCTGAACTTTCTGAAGCAAC
SiPUB46-F	TCAGAGAAATTGACGAGGAG
SiPUB46-R	ATAATCCTGTCAGCAGGTTG
SiPUB49-F	GGAGCGAGATGATGCGGTTA
SiPUB49-R	AGTGAACGGCTGAAGACCTG
SiPUB55-F	CCAGCATCGAGTCCTGGGTC
SiPUB55-R	ACGCACCACTCCTGGATGA
SiPUB59-F	CAACAAGCCCAAGAGGGACT
SiPUB59-R	GCTGGCTTCCCAGTTACCAT
SiPUB64-F	CTCACAGGAACAAAGCGTGC
SiPUB64-R	CACGCCTCAATGTGCTGTTC
SiPUB70-F	CTGTTCCTGTGCCCCATCTC
SiPUB70-R	CCGTACCCGAAGATCCACTG

分类名称	蛋白名称	基因
Classification name	Protein name	Gene
Ⅰ	U-box+ARM	SiPUB5， SiPUB10， SiPUB12， SiPUB14， SiPUB25， SiPUB31， SiPUB32， SiPUB33， SiPUB35， SiPUB37， SiPUB43， SiPUB47， SiPUB48， SiPUB64， SiPUB65， SiPUB71
Ⅱ	U-box	SiPUB2， SiPUB15， SiPUB16， SiPUB19， SiPUB20， SiPUB21， SiPUB22， SiPUB26， SiPUB27， SiPUB34， SiPUB40， SiPUB42， SiPUB45， SiPUB55， SiPUB57， SiPUB62， SiPUB66， SiPUB67， SiPUB70
Ⅲ	U-box	SiPUB17， SiPUB18， SiPUB58
Ⅲ	U-box+KAP	SiPUB50
Ⅳ	U-box	SiPUB28
Ⅳ	U-box+ARM	SiPUB9， SiPUB28， SiPUB54
Ⅴ	U-box	SiPUB13， SiPUB23， SiPUB56
	U-box+ARM	SiPUB39
	U-box+HEAT_2	SiPUB3
Ⅵ	U-box	SiPUB8
Ⅵ	Pro_isomerase+U-box	SiPUB44
Ⅶ	U-box	SiPUB6， SiPUB61
	U-box+Pkinase	SiPUB1， SiPUB36， SiPUB38， SiPUB46， SiPUB52， SiPUB68
	U-box+Pkinase+Usp	SiPUB7， SiPUB59， SiPUB63
	U-box+Pkinase_Tyr+Usp	SiPUB11， SiPUB41， SiPUB51， SiPUB69
	TPR_11+U-box+Pkinase	SiPUB30， SiPUB60
Ⅸ	Prp19+WD40+U-box	SiPUB29

分类名称	蛋白名称	基因
Classification name	Protein name	Gene
Ⅰ	U-box+ARM	SiPUB5， SiPUB10， SiPUB12， SiPUB14， SiPUB25， SiPUB31， SiPUB32， SiPUB33， SiPUB35， SiPUB37， SiPUB43， SiPUB47， SiPUB48， SiPUB64， SiPUB65， SiPUB71
Ⅱ	U-box	SiPUB2， SiPUB15， SiPUB16， SiPUB19， SiPUB20， SiPUB21， SiPUB22， SiPUB26， SiPUB27， SiPUB34， SiPUB40， SiPUB42， SiPUB45， SiPUB55， SiPUB57， SiPUB62， SiPUB66， SiPUB67， SiPUB70
Ⅲ	U-box	SiPUB17， SiPUB18， SiPUB58
Ⅲ	U-box+KAP	SiPUB50
Ⅳ	U-box	SiPUB28
Ⅳ	U-box+ARM	SiPUB9， SiPUB28， SiPUB54
Ⅴ	U-box	SiPUB13， SiPUB23， SiPUB56
	U-box+ARM	SiPUB39
	U-box+HEAT_2	SiPUB3
Ⅵ	U-box	SiPUB8
Ⅵ	Pro_isomerase+U-box	SiPUB44
Ⅶ	U-box	SiPUB6， SiPUB61
	U-box+Pkinase	SiPUB1， SiPUB36， SiPUB38， SiPUB46， SiPUB52， SiPUB68
	U-box+Pkinase+Usp	SiPUB7， SiPUB59， SiPUB63
	U-box+Pkinase_Tyr+Usp	SiPUB11， SiPUB41， SiPUB51， SiPUB69
	TPR_11+U-box+Pkinase	SiPUB30， SiPUB60
Ⅸ	Prp19+WD40+U-box	SiPUB29

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