Analysis of the Potato SUMO E3 Ligase Gene Family and Cloning and Expression Pattern of StSIZ1

doi:10.13560/j.cnki.biotech.bull.1985.2024-1252

Abstract

Abstract:

Objective This study is aimed to clone, characterize, and analyze the subcellular localization and tissue-specific expression of the StSIZ1 in potato, providing theoretical foundation for elucidating its functional role. Method Homology search was conducted using the Arabidopsis thaliana SUMO E3 ligase gene sequence to obtain members of the potato SUMO E3 ligase gene family, and have bioinformatics analysis of them. The StSIZ1 was cloned from the potato variety Atlantic using reverse transcription PCR (RT-PCR). The expression patterns of StSIZ1 in various potato tissues and its responses to abiotic stresses and hormone were analyzed using quantitative real-time PCR (RT-qPCR). A pCEGFP-StSIZ1 subcellular localization vector was constructed, and the localization of StSIZ1 was confirmed via Agrobacterium-mediated transient expression in tobacco. Result The coding sequence (CDS) of the StSIZ1 is 2 634 base pairs (bp) in length and is located on chromosome 11. RT-qPCR results revealed that StSIZ1 has the lowest relative expression in stems and the highest in tubers. The gene presents significant responsiveness to various abiotic stress treatments, including osmotic, salt, and high-temperature stress. Subcellular localization analysis using fluorescence detection indicated that StSIZ1 primarily functions within the nucleus. Conclusion The StSIZ1 is responsive to a wide range of abiotic stresses and functions in the nucleus, highlighting its potential role in tolerance to stress.

Key words: potato, StSIZ1, gene cloning, subcellular localization, expression pattern analysis

XU Hui-zhen, SHANTWANA Ghimire, RAJU Kharel, YUE Yun, SI Huai-jun, TANG Xun. Analysis of the Potato SUMO E3 Ligase Gene Family and Cloning and Expression Pattern of StSIZ1[J]. Biotechnology Bulletin, 2025, 41(6): 119-129.

Figures/Tables 9

References 39

1	Scaramagli S, Biondi S, Leone A, et al. Acclimation to low water potential in potato cell suspension cultures leads to changes in putrescine metabolism [J]. Plant Physiol Biochem, 2000, 38(4): 345-351.
2	贾明飞, 樊建英, 封志明, 等. 氮素不同形态对早熟马铃薯产量和氮素积累的影响 [J]. 中国土壤与肥料, 2024, (2): 146-151.
	Jia MF, Fan JY, Feng ZM, et al. Effects of different forms of nitrogen on yield and nitrogen accumulation in early maturing potatoes [J]. Soil and Fertiliser China, 2024, (2): 146-151.
3	汪昕, 杨德秋, 李洋, 等. 马铃薯机械化收获技术进展与展望 [J]. 农机化研究, 2024, 46(9): 1-7.
	Wang X, Yang DQ, Li Y, et al. Progress and prospect of potato mechanized harvesting technology [J]. J Agric Mech Res, 2024, 46(9): 1-7.
4	Seo J, Lee KJ. Post-translational modifications and their biological functions: proteomic analysis and systematic approaches [J]. J Biochem Mol Biol, 2004, 37(1): 35-44.
5	Yeh ET, Gong L, Kamitani T. Ubiquitin-like proteins: new wines in new bottles [J]. Gene, 2000, 248(1/2): 1-14.
6	Kurepa J, Walker JM, Smalle J, et al. The small ubiquitin-like modifier (SUMO) protein modification system in Arabidopsis. Accumulation of SUMO1 and-2 conjugates is increased by stress [J]. J Biol Chem, 2003, 278(9): 6862-6872.
7	Elrouby N. Analysis of small ubiquitin-like modifier (SUMO) targets reflects the essential nature of protein SUMOylation and provides insight to elucidate the role of SUMO in plant development [J]. Plant Physiol, 2015, 169(2): 1006-1017.
8	Park HJ, Yun DJ. SUMO proteins grapple with biotic and abiotic stresses in Arabidopsis [J]. J Plant Biol, 2013, 56(2): 77-84.
9	Novatchkova M, Tomanov K, Hofmann K, et al. Update on sumoylation: defining core components of the plant SUMO conjugation system by phylogenetic comparison [J]. New Phytol, 2012, 195(1): 23-31.
10	Han YF, Zhao QY, Dang LL, et al. The SUMO E3 ligase-like proteins PIAL1 and PIAL2 interact with MOM1 and form a novel complex required for transcriptional silencing [J]. Plant Cell, 2016, 28(5): 1215-1229.
11	Ritterhoff T, Das H, Hofhaus G, et al. The RanBP2/RanGAP1*SUMO1/Ubc9 SUMO E3 ligase is a disassembly machine for Crm1-dependent nuclear export complexes [J]. Nat Commun, 2016, 7: 11482.
12	Zheng Z, Liu D. SIZ1 regulates phosphate deficiency-induced inhibition of primary root growth of Arabidopsis by modulating Fe accumulation and ROS production in its roots [J]. Plant Signal Behav, 2021, 16(10): 1946921.
13	Rawat SS, Sandhya S, Laxmi A. Complex genetic interaction between glucose sensor HXK1 and E3 SUMO ligase SIZ1 in regulating plant morphogenesis [J]. Plant Signal Behav, 2024, 19(1): 2341506.
14	Gao SJ, Zeng XQ, Wang JH, et al. Arabidopsis SUMO E3 ligase SIZ1 interacts with HDA6 and negatively regulates HDA6 function during flowering [J]. Cells, 2021, 10(11): 3001.
15	Castro PH, Couto D, Santos MÂ, et al. SUMO E3 ligase SIZ1 connects sumoylation and reactive oxygen species homeostasis processes in Arabidopsis [J]. Plant Physiol, 2022, 189(2): 934-954.
16	Miura K, Nozawa R. Overexpression of SIZ1 enhances tolerance to cold and salt stresses and attenuates response to abscisic acid in Arabidopsis thaliana [J]. Plant Biotechnol, 2014, 31(2): 167-172.
17	Kim JY, Song JT, Seo HS. Post-translational modifications of Arabidopsis E3 SUMO ligase AtSIZ1 are controlled by environmental conditions [J]. FEBS Open Bio, 2017, 7(10): 1622-1634.
18	Yoo CY, Miura K, Jin JB, et al. SIZ1 small ubiquitin-like modifier E3 ligase facilitates basal thermotolerance in Arabidopsis independent of salicylic acid [J]. Plant Physiol, 2006, 142(4): 1548-1558.
19	Miura K, Sato A, Ohta M, et al. Increased tolerance to salt stress in the phosphate-accumulating Arabidopsis mutants siz1 and pho2 [J]. Planta, 2011, 234(6): 1191-1199.
20	Jiang H, Zhou LJ, Gao HN, et al. The transcription factor MdMYB2 influences cold tolerance and anthocyanin accumulation by activating SUMO E3 ligase MdSIZ1 in apple [J]. Plant Physiol, 2022, 189(4): 2044-2060.
21	张亚丽. 苹果SUMO E3连接酶MdSIZ1蛋白SUMO化MdMYB30调控蜡质形成的机理 [D]. 泰安: 山东农业大学, 2023.
	Zhang YL. Mechanism of regulating wax formation by sumoylation of apple SUMO E3 ligase MdSIZ1 protein MdMYB30 [D]. Tai'an: Shandong Agricultural University, 2023.
22	Zhang S, Wang SJ, Lv JL, et al. SUMO E3 ligase SlSIZ1 facilitates heat tolerance in tomato [J]. Plant Cell Physiol, 2018, 59(1): 58-71.
23	Zhang S, Zhuang KY, Wang SJ, et al. A novel tomato SUMO E3 ligase, SlSIZ1, confers drought tolerance in transgenic tobacco [J]. J Integr Plant Biol, 2017, 59(2): 102-117.
24	Cai B, Kong XX, Zhong C, et al. SUMO E3 Ligases GmSIZ1a and GmSIZ1b regulate vegetative growth in soybean [J]. J Integr Plant Biol, 2017, 59(1): 2-14.
25	Li YJ, Wang GX, Xu ZQ, et al. Organization and regulation of soybean SUMOylation system under abiotic stress conditions [J]. Front Plant Sci, 2017, 8: 1458.
26	Lei SK, Wang QZ, Chen Y, et al. Capsicum SIZ1 contributes to ABA-induced SUMOylation in pepper [J]. Plant Sci, 2022, 314: 111099.
27	Lai RQ, Jiang JM, Wang J, et al. Functional characterization of three maize SIZ/PIAS-type SUMO E3 ligases [J]. J Plant Physiol, 2022, 268: 153588.
28	汪永平. 梭梭SUMO E3连接酶基因HaSIZ1的克隆和功能分析 [D]. 兰州: 兰州大学, 2020.
	Wang YP. Cloning and functional analysis of HaSIZ1 ligase gene of Haloxylon ammodendron SUMO E3 [D]. Lanzhou: Lanzhou University, 2020.
29	Kumar S, Stecher G, Tamura K. MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets [J]. Mol Biol Evol, 2016, 33(7): 1870-1874.
30	Chen CJ, Wu Y, Li JW, et al. TBtools-II: a "one for all, all for one" bioinformatics platform for biological big-data mining [J]. Mol Plant, 2023, 16(11): 1733-1742.
31	Srivastava M, Verma V, Srivastava AK. The converging path of protein SUMOylation in phytohormone signalling: highlights and new frontiers [J]. Plant Cell Rep, 2021, 40(11): 2047-2061.
32	Crozet P, Margalha L, Butowt R, et al. SUMOylation represses SnRK1 signaling in Arabidopsis [J]. Plant J, 2016, 85(1): 120-133.
33	Xiong JW, Yang FB, Wei F, et al. Inhibition of SIZ1-mediated SUMOylation of HOOKLESS1 promotes light-induced apical hook opening in Arabidopsis [J]. Plant Cell, 2023, 35(6): 2027-2043.
34	Cohen-Peer R, Schuster S, Meiri D, et al. Sumoylation of Arabidopsis heat shock factor A2 (HsfA2) modifies its activity during acquired thermotholerance [J]. Plant Mol Biol, 2010, 74(1/2): 33-45.
35	Garcia-Dominguez M, March-Diaz R, Reyes JC. The PHD domain of plant PIAS proteins mediates sumoylation of bromodomain GTE proteins [J]. J Biol Chem, 2008, 283(31): 21469-21477.
36	Suzuki R, Shindo H, Tase A, et al. Solution structures and DNA binding properties of the N-terminal SAP domains of SUMO E3 ligases from Saccharomyces cerevisiae and Oryza sativa [J]. Proteins, 2009, 75(2): 336-347.
37	Catala R, Ouyang J, Abreu IA, et al. The Arabidopsis E3 SUMO ligase SIZ1 regulates plant growth and drought responses [J]. Plant Cell, 2007, 19(9): 2952-2966.
38	赵媛媛, 刘萍涛, 康建宏. 高温胁迫对马铃薯叶片生理特性及产量影响研究 [J]. 中国农学通报, 2024, 40(21): 35-44.
	Zhao YY, Liu PT, Kang JH. Effects of high temperature stress on physiological characteristics and yield of potato leaves [J]. Chin Agric Sci Bull, 2024, 40(21): 35-44.
39	Wyrzykowska A, Bielewicz D, Plewka P, et al. The MYB33, MYB₆5, and MYB101 transcription factors affect Arabidopsis and potato responses to drought by regulating the ABA signaling pathway [J]. Physiol Plant, 2022, 174(5): e13775.

引物名称 Primer name	引物序列 Primer sequence （5′-3′）	用途 Purpose
pCAMBIA1300-StSIZ1-F	ggtacccggggatcctctagaATGGATTTGGTTGCTAGCTGCA	基因克隆
pCAMBIA1300-StSIZ1-R	gcccttgctcaccatgtcgacCTATTCAGAATCCGAATCAATACTTAGAT	基因克隆
qPCR-StSIZ1-F	TTAGACTTGTGAAGCGGCGT	RT-qPCR
qPCR-StSIZ1-R	CCACCAACACAACGACGAAC	RT-qPCR
Eflα-F	GATGGTCAGACCCGTGAACA	内参基因
Eflα-R	CCTTGGAGTACTTCGGGGTC	内参基因

引物名称 Primer name	引物序列 Primer sequence （5′-3′）	用途 Purpose
pCAMBIA1300-StSIZ1-F	ggtacccggggatcctctagaATGGATTTGGTTGCTAGCTGCA	基因克隆
pCAMBIA1300-StSIZ1-R	gcccttgctcaccatgtcgacCTATTCAGAATCCGAATCAATACTTAGAT	基因克隆
qPCR-StSIZ1-F	TTAGACTTGTGAAGCGGCGT	RT-qPCR
qPCR-StSIZ1-R	CCACCAACACAACGACGAAC	RT-qPCR
Eflα-F	GATGGTCAGACCCGTGAACA	内参基因
Eflα-R	CCTTGGAGTACTTCGGGGTC	内参基因

基因ID Gene ID	基因名 Gene name	理论分子量 Molecular weight （Da）	等电点 Point isoelectric	不稳定指数 Instability index	脂肪族氨基酸指数 Aliphatic index	亲水度 Grand average of hydropathicity
Soltu.DM.11G022540.5	StSIZ1	95 449.87	4.96	44.28	76.34	-0.453
Soltu.DM.06G005690.1	StSIZ2	93 832.16	5.44	47.05	80.67	-0.373
Soltu.DM.07G023970.1	StMMS21	28 532.31	5.10	58.06	77.34	-0.533
Soltu.DM.08G004520.1	StPIAS1	96 099.04	6.01	48.14	73.54	-0.390
Soltu.DM.06G005850.1	StPIAS2	163 534.42	6.74	41.34	88.45	-0.296
Soltu.DM.05G011900.1	StRanBP2a	30 458.93	9.24	58.23	29.93	-0.922
Soltu.DM.12G024390.1	StRanBP2b	26 349.84	8.17	52.64	58.67	-0.482
Soltu.DM.09G019740.1	StRanGAP1a	58 094.47	4.58	37.95	94.34	-0.283
Soltu.DM.01G024550.1	StRanGAP1b	60 594.10	4.64	41.86	90.05	-0.386
Soltu.DM.05G007610.1	StRanGAP1c	64 532.83	8.78	30.85	99.40	-0.067

基因ID Gene ID	基因名 Gene name	理论分子量 Molecular weight （Da）	等电点 Point isoelectric	不稳定指数 Instability index	脂肪族氨基酸指数 Aliphatic index	亲水度 Grand average of hydropathicity
Soltu.DM.11G022540.5	StSIZ1	95 449.87	4.96	44.28	76.34	-0.453
Soltu.DM.06G005690.1	StSIZ2	93 832.16	5.44	47.05	80.67	-0.373
Soltu.DM.07G023970.1	StMMS21	28 532.31	5.10	58.06	77.34	-0.533
Soltu.DM.08G004520.1	StPIAS1	96 099.04	6.01	48.14	73.54	-0.390
Soltu.DM.06G005850.1	StPIAS2	163 534.42	6.74	41.34	88.45	-0.296
Soltu.DM.05G011900.1	StRanBP2a	30 458.93	9.24	58.23	29.93	-0.922
Soltu.DM.12G024390.1	StRanBP2b	26 349.84	8.17	52.64	58.67	-0.482
Soltu.DM.09G019740.1	StRanGAP1a	58 094.47	4.58	37.95	94.34	-0.283
Soltu.DM.01G024550.1	StRanGAP1b	60 594.10	4.64	41.86	90.05	-0.386
Soltu.DM.05G007610.1	StRanGAP1c	64 532.83	8.78	30.85	99.40	-0.067

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