Please wait a minute...
Journal of Integrative Agriculture  2020, Vol. 19 Issue (9): 2239-2246    DOI: 10.1016/S2095-3119(20)63262-2
Special Issue: 园艺-分子生物合辑Horticulture — Genetics · Breeding
Horticulture Advanced Online Publication | Current Issue | Archive | Adv Search |
Overexpression of StCYS1 gene enhances tolerance to salt stress in the transgenic potato (Solanum tuberosum L.) plant
LIU Min-min1, LI Ya-lun1, LI Guang-cun2, DONG Tian-tian1, LIU Shi-yang1, LIU Pei1, WANG Qing-guo1
1 Key Laboratory of Food Processing Technology and Quality Control in Shandong Province, College of Food Science and Engineering, Shandong Agricultural University, Tai’an 271018, P.R.China
2 Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences/Key Laboratory of Biology and Genetic Improvement of Tuber and Root Crop, Ministry of Agriculture and Rural Affairs, Beijing 100081, P.R.China
Download:  PDF in ScienceDirect  
Export:  BibTeX | EndNote (RIS)      
Salt stress seriously restricts the growth and yield of potatoes.  Plant cystatins are vital players in biotic stress and development, however, their roles in salt stress resistance remain elusive.  Here, we report that StCYS1 positively regulates salt tolerance in potato plants.  An in vitro biochemical test demonstrated that StCYS1 is a bona fide cystatin.  Overexpression of StCYS1 in both Escherichia coli and potato plants significantly increased their resistance to high salinity.  Further analysis revealed that the transgenic plants accumulated more proline and chlorophyll under salt stress conditions.  Moreover, the transgenic plants displayed higher H2O2 scavenging capability and cell membrane integrity compared with wild-type potato.  These results demonstrate that StCYS1 is closely correlated with salt stress and its overaccumulation can substantially enhance salt stress resistance.
Keywords:  potato        salt stress        cystatin  StCYS1       overexpression  
Received: 06 January 2020   Accepted:
Fund: This work was supported by the National Natural Science Foundation of China (31901752) and by a grant from the Potato Industry Innovation Team for Modern Agricultural Industry Technology System, Shandong Province, China (SDAIT-10-011-11).
Corresponding Authors:  Correspondence LIU Pei, Tel/Fax: +86-538-8246021, E-mail:; WANG Qing-guo, E-mail:   
About author:  LIU Min-min, E-mail:;

Cite this article: 

LIU Min-min, LI Ya-lun, LI Guang-cun, DONG Tian-tian, LIU Shi-yang, LIU Pei, WANG Qing-guo. 2020. Overexpression of StCYS1 gene enhances tolerance to salt stress in the transgenic potato (Solanum tuberosum L.) plant. Journal of Integrative Agriculture, 19(9): 2239-2246.

Benchabane M, Schlüter U, Vorster J, Goulet M C, Michaud D. 2010. Plant cystatins. Biochimie, 92, 1657–1666.
Birch P R J, Bryan G, Fenton B, Gilroy E M, Hein I, Jones J T, Prashar A, Taylor M A, Torrance L, Toth I K. 2012. Crops that feed the world 8: Potato: Are the trends of increased global production sustainable? Food Security, 4, 477–508.
Bruce M A, Shoup Rupp J L. 2019. Agrobacterium-mediated transformation of Solanum tuberosum L., potato. In: Kumar S, Barone P, Smith M, eds. Transgenic Plants. Methods in Molecular Biology. vol. 1864. Humana Press, New York, NY.
Chen G Q, Zhang D, Shen X H. 2018. Cloning and characterization of ApCystatin, a plant cystatin gene from Agapanthus praecox ssp. orientalis responds to abiotic stress. Protein Expression and Purification, 149, 66–74.
Christova P K, Christov N K, Mladenov P V, lmai R. 2018. The wheat multidomain cystatin TaMDC1 displays antifungal, antibacterial, and insecticidal activities in planta. Plant Cell Reports, 37, 923–932.
Cingel A, Savi? J, Lazarevi? J, Cosi? T, Raspor M, Smigocki A, Ninkovi? S. 2017. Co-expression of the proteinase inhibitors oryzacystatin I and oryzacystatin II in transgenic potato alters Colorado potato beetle larval development. Insect Science, 24, 768–780.
Cingel A, Savi? J, Vinterhalter B, Vinterhalter D, Kosti? M, Šešlija Jovanovi? D, Smigocki A, Ninkovi? S. 2015. Growth and development of Colorado potato beetle larvae, Leptinotarsa decemlineata, on potato plants expressing the oryzacystatin II proteinase inhibitor. Transgenic Research, 24, 729–740.
Deinlein U, Stephan A B, Horie T, Luo W, Xu G, Schroeder J I. 2014. Plant salt-tolerance mechanisms. Trends in Plant Science, 19, 371–379.
El-Banna A, Taller J. 2017. Functional characterization of the silenced potato cysteine proteinase inhibitor gene (PCPI) in Phytophthora infestans resistance. Physiological and Molecular Plant Pathology, 100, 23–29.
Gaddour K, Vicente-Carbajosa J, Lara P, Isabel-Lamoneda I, Díaz I, Carbonero P. 2001. A constitutive cystatin-encoding gene from barley (Icy) responds differentially to abiotic stimuli. Plant Molecular Biology, 45, 599–608.
Gangadhar B H, Sajeesh K, Venkatesh J, Baskar V, Abhinandan K, Yu J W, Prasad R, Mishra R K. 2016. Enhanced tolerance of transgenic potato plants over-expressing non-specific lipid transfer protein-1 (StnsLTP1) against multiple abiotic stresses. Frontiers in Plant Science, 7, 1228.
Girard C, Rivard D, Kiggundu A, Kunert K, Gleddie S C, Cloutier C, Michaud D. 2007. A multicomponent, elicitor inducible cystatin complex in tomato, Solanum lycopersicum. New Phytologist, 173, 841–851.
Green A R, Nissen M S, Kumar G N M, Knowles N R, Kang C. 2013. Characterization of Solanum tuberosum multicystatin and the significance of core domains. The Plant Cell, 25, 5043–5052.
He Y, Yang X, Xu C, Guo D, Niu L, Wang Y, Li J, Yan F, Wang Q. 2018. Overexpression of a novel transcriptional repressor GmMYB3a negatively regulates salt-alkali tolerance and stress-related genes in soybean. Biochemical and Biophysical Research Communications, 498, 586–591.
Hemavathi, Upadhyaya C P, Young K E, Akula N, Kim H S, Heung J J, Oh O M, Aswath C R, Chun S C, Kim D H, Park S W. 2009. Over-expression of strawberry D-galacturonic acid reductase in potato leads to accumulation of vitamin C with enhanced abiotic stress tolerance. Plant Science, 177, 659–667.
Isayenkov S V, Maathuis F J M. 2019. Plant salinity stress: Many unanswered questions remain. Frontiers in Plant Science, 10, 80.
Jaarsma R, de Boer A H. 2018. Salinity tolerance of two potato cultivars (Solanum tuberosum) correlates with differences in vacuolar transport activity. Frontiers in Plant Science, 9, 737.
Jaarsma R, de Vries R S M, de Boer A H. 2013. Effect of salt stress on growth, Na+ accumulation and proline metabolism in potato (Solanum tuberosum) cultivars. PLoS ONE, 8, e60183.
Jia F, Wan X, Zhu W, Sun D, Zheng C, Liu P, Huang J. 2015. Overexpression of mitochondrial phosphate transporter 3 severely hampers plant development through regulating mitochondrial function in Arabidopsis. PLoS ONE, 10, e0129717.
Li X, Guo C, Ahmad S, Wang Q, Yu J, Liu C, Guo Y. 2019. Systematic analysis of MYB family genes in potato and their multiple roles in development and stress responses. Biomolecules, 9, 317.
Liang W, Ma X, Wan P, Liu L. 2018. Plant salt-tolerance mechanism: A review. Biochemical and Biophysical Research Communications, 495, 286–291.
Lival K J, Schmittgen T D. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the the 2–ΔΔCT method. Methods, 25, 402–408.
Ma Y R, Yang M N, Wang J J, Jiang C Z, Wang Q G. 2017.  Application of exogenous ethylene inhibits postharvest peel browning of ‘Huangguan’ pear. Frontiers in Plant Science, 7, 2029.
Massonneau A, Condamine P, Wisniewski J P, Zivy M, Rogowsky P M. 2005. Maize cystatins respond to developmental cues, cold stress and drought. Biochimica et Biophysica Acta, 1729, 186–199.
Muchate N S, Nikalje G C, Rajurkar N S, Suprasanna P, Nikam T D. 2016. Plant salt stress: Adaptive responses, tolerance mechanism and bioengineering for salt tolerance. The Botanical Review, 82, 371–406.
Munger A, Coenen K, Cantin L, Goulet C, Vaillancourt L P, Goulet M C, Tweddell R, Sainsbury F, Michaud D. 2012. Beneficial ‘unintended effects’ of a cereal cystatin in transgenic lines of potato, Solanum tuberosum. BMC Plant Biology, 12, 198.
Munger A, Simon M A, Khalf M, Goulet M C, Michaud D. 2015. Cereal cystatins delay sprouting and nutrient loss in tubers of potato, Solanum tuberosum. BMC Plant Biology, 15, 296.
Nissen M S, Kumar G N M, Youn B, Knowles D B, Lam K S, Ballinger W J, Knowles N R, Kang C. 2009. Characterization of Solanum tuberosum multicystatin and its structural comparison with other cystatins. The Plant Cell, 21, 861–875.
Shi J, Wang J, Wang N, Zhou H, Xu Q, Yan G. 2019. Overexpression of StGA2ox1 gene increases the tolerance to abiotic stress in transgenic potato (Solanum tuberosum L.) plants. Applied Biochemistry and Biotechnology, 187, 1204–1219.
Shyu D J H, Chou W M, Yiu T J, Lin C P C, Tzen J T C. 2004. Cloning, functional expression, and characterization of cystatin in sesame seed. Journal of Agricultural and Food Chemistry, 52, 1350–1356.
Silveira J A G, Viégas R A, da Rocha I M A, Moreira A C O M, Moreira R A, Oliveira J T A. 2003. Proline accumulation and glutamine synthetase activity are increased by salt-induced proteolysis in cashew leaves. Journal of Plant Physiology, 160, 115–123.
Šmid I, Gruden K, Gašpari? M B, Koruza K, Petek M, Pohleven J, Brzin J, Kos J, ?el J, Saboti? J. 2013. Inhibition of the growth of Colorado potato beetle larvae by macrocypins, protease inhibitors from the parasol mushroom. Journal of Agricultural and Food Chemistry, 61, 12499–12509.
Šmid I, Rotter A, Gruden K, Brzin J, Gašpari? M B, Kos J, ?el J, Saboti? J. 2015. Clitocypin, a fungal cysteine protease inhibitor, exerts its insecticidal effect on Colorado potato beetle larvae by inhibiting their digestive cysteine proteases. Pesticide Biochemistry and Physiology, 122, 59–66.
Tan Y, Li M, Yang Y, Sun X, Wang N, Liang B, Ma F. 2017. Overexpression of MpCYS4, a phytocystatin gene from Malus prunifolia (Willd.) Borkh., enhances stomatal closure to confer drought tolerance in transgenic Arabidopsis and apple. Frontiers in Plant Science, 8, 33.
Tiwari L A, Mittal D, Mishra R C, Grover A. 2015. Constitutive over-expression of rice chymotrypsin protease inhibitor gene OCPI2 results in enhanced growth, salinity and osmotic stress tolerance of the transgenic Arabidopsis plants. Plant Physiology and Biochemistry, 92, 48–55.
Tremblay J, Goulet M C, Michaud D. 2019. Recombinant cystatins in plants. Biochimie, 166, 184–193.
Waldron C, Wegrich L M, Merlo P A O, Walsh T A. 1993. Characterization of a genomic sequence coding for potato multicystatin, an eight-domain cysteine proteinase inhibitor. Plant Molecular Biology, 23, 801–812.
Wang L, Liu Y, Li D, Feng S, Yang J, Zhang J, Zhang J, Wang D, Gan Y. 2019. Improving salt tolerance in potato through overexpression of AtHKT1 gene. BMC Plant Biology, 19, 357.
Wang W, Zhou X M, Xiong H X, Mao W Y, Zhao P, Sun M X. 2018. Papain-like and legumain-like proteases in rice: genome-wide identification, comprehensive gene feature characterization and expression analysis. BMC Plant Biology, 18, 87.
Wang Y, Zhan Y, Wu C, Gong S, Zhu N,Chen S, Li H. 2012. Cloning of a cystatin gene from sugar beet M14 that can enhance plant salt tolerance. Plant Science, 191–192, 93–99.
Wu J, Haard N F. 2000. Purification and characterization of a cystatin from the leaves of methyl jasmonate treated tomato plants. Comparative Biochemistry and Physiology Part C: Pharmacology, Toxicology and Endocrinology, 127, 209–220.
Van Wyk S G, Kunert K J, Cullis C A, Pillay P, Makgopa M E, Schlüter U, Vorster B J. 2016. Review: The future of cystatin engineering. Plant Science, 246, 119–127.
Yang Y, Guo Y. 2018. Elucidating the molecular mechanisms mediating plant salt-stress responses. New Phytologist, 217, 523–539.
You L, Song Q, Wu Y, Li S, Jiang C, Chang L, Yang X, Zhang J. 2019. Accumulation of glycine betaine in transplastomic potato plants expressing choline oxidase confers improved drought tolerance. Planta, 249, 1963–1975.
Yu Z F, Cao J X, Zhu S H, Zhang L L, Peng Y, Shi J Y. 2020. Exogenous nitric oxide enhances disease resistance by nitrosylation and inhibition of S-nitrosoglutathione reductase in peach fruit. Frontiers in Plant Science, 11, 543.
Zhang X, Liu S, Takano T. 2008. Two cysteine proteinase inhibitors from Arabidopsis thaliana, AtCYSa and AtCYSb, increasing the salt, drought, oxidation and cold tolerance. Plant Molecular Biology, 68, 131–143.
Zhao Y, Botella M A, Subramanian L, Niu X, Nielsen S S, Bressan R A, Hasegawa P M. 1996. Two wound-inducible soybean cysteine proteinase inhibitors have greater insect digestive proteinase inhibitory activities than a constitutive homolog. Plant Physiology, 111, 1299–1306.
Zhou X, Zhang N, Yang J, Si H. 2016. Functional analysis of potato CPD gene: A rate-limiting enzyme in brassinosteroid biosynthesis under polyethylene glycol-Induced osmotic stress. Crop Science, 56, 2675–2687.
Zhu W, Bai X, Li G, Chen M, Wang Z, Yang Q. 2019. SpCYS, a cystatin gene from wild potato (Solanum pinnatisectum), is involved in the resistance against Spodoptera litura. Theoretical and Experimental Plant Physiology, 31, 317–328.
[1] XIAO Yang-yang, QIAN Jia-jia, HOU Xing-liang, ZENG Lan-ting, LIU Xu, MEI Guo-guo, LIAO Yin-yin.

Diurnal emission of herbivore-induced (Z)-3-hexenyl acetate and allo-ocimene activates sweet potato defense responses to sweet potato weevils [J]. >Journal of Integrative Agriculture, 2023, 22(6): 1782-1796.

[2] MA Xiao-wen, MA Qiu-xiang, MA Mu-qing, CHEN Yan-hang, GU Jin-bao, LI Yang, HU Qing, LUO Qing-wen, WEN Ming-fu, ZHANG Peng, LI Cong, WANG Zhen-yu.

Cassava MeRS40 is required for the regulation of plant salt tolerance [J]. >Journal of Integrative Agriculture, 2023, 22(5): 1396-1411.

[3] AI Ju, WANG Ye, YAN Ya-wen, LI Chen-xiao, LUO Wei, MA Ling, SHANG Yi, GAO Dong-li. StOFP20 regulates tuber shape and interacts with TONNEAU1 Recruiting Motif proteins in potato[J]. >Journal of Integrative Agriculture, 2023, 22(3): 752-761.
[4] LI Rui-jie, ZHAI Hong, HE Shao-zhen, ZHANG Huan, ZHAO Ning, LIU Qing-chang. A geranylgeranyl pyrophosphate synthase gene, IbGGPS, increases carotenoid contents in transgenic sweetpotato[J]. >Journal of Integrative Agriculture, 2022, 21(9): 2538-2546.
[5] WANG Chu-kun, ZHAO Yu-wen, HAN Peng-liang, YU Jian-qiang, HAO Yu-jin, XU Qian, YOU Chun-xiang, HU Da-gang. Auxin response factor gene MdARF2 is involved in ABA signaling and salt stress response in apple[J]. >Journal of Integrative Agriculture, 2022, 21(8): 2264-2274.
[6] YANG Sheng-di, GUO Da-long, PEI Mao-song, WEI Tong-lu, LIU Hai-nan, BIAN Lu, YU Ke-ke, ZHANG Guo-hai, YU Yi-he. Identification of the DEAD-box RNA helicase family members in grapevine reveals that VviDEADRH25a confers tolerance to drought stress[J]. >Journal of Integrative Agriculture, 2022, 21(5): 1357-1374.
[7] ZHU Yu, YUAN Yu-han, MEI Li-ping, DING Shuang-kun, GAO Yu-chen, DU Xian-feng, GUO Li. Comparison of structural and physicochemical properties of potato protein and potato flour modified with tyrosinase[J]. >Journal of Integrative Agriculture, 2022, 21(5): 1513-1524.
[8] DONG Suo-meng, ZHOU Shao-qun. Potato late blight caused by Phytophthora infestans: From molecular interactions to integrated management strategies[J]. >Journal of Integrative Agriculture, 2022, 21(12): 3456-3466.
[9] LI Chen, LIU Xuan-xuan, ABOUELNASR Hesham, MOHAMED HAMED Arisha, KOU Meng, TANG Wei, YAN Hui, WANG Xin, WANG Xiao-xiao, ZHANG Yun-gang, LIU Ya-ju, GAO Run-fei, MA Meng, LI Qiang. Inhibition of miR397 by STTM technology to increase sweetpotato resistance to SPVD[J]. >Journal of Integrative Agriculture, 2022, 21(10): 2865-2875.
[10] LI Yan-yan, GUO Li-na, LIANG Cheng-zhen, MENG Zhi-gang, Syed Tahira, GUO San-dui, ZHANG Rui. Overexpression of Brassica napus cytosolic fructose-1,6-bisphosphatase and sedoheptulose-1,7-bisphosphatase genes significantly enhanced tobacco growth and biomass[J]. >Journal of Integrative Agriculture, 2022, 21(1): 49-59.
[11] WANG Yun-shu, GUO Peng-yu, ZHANG Jian-ling, XIE Qiao-li, SHEN Hui, HU Zong-li, CHEN Guo-ping. Overexpression of the MADS-box gene SlMBP21 alters leaf morphology and affects reproductive development in tomato[J]. >Journal of Integrative Agriculture, 2021, 20(12): 3170-3185.
[12] CHEN Li-li, WANG Hao-ying, GONG Xiao-chen, ZENG Zhao-hai, XUE Xu-zhang, HU Yue-gao. Transcriptome analysis reveals effects of red and blue lightemitting diodes (LEDs) on the growth, chlorophyll fluorescence and endogenous plant hormones of potato (Solanum tuberosum L.) plantlets cultured in vitro[J]. >Journal of Integrative Agriculture, 2021, 20(11): 2914-2931.
[13] WU Jia-yu, ZHANG Yu, ZHOU Xue-ping, QIAN Ya-juan. Three sensitive and reliable serological assays for detection of potato virus A in potato plants[J]. >Journal of Integrative Agriculture, 2021, 20(11): 2966-2975.
[14] YIN Jian, CHENG Li, HONG Yan, LI Zhao-feng, LI Cai-ming, BAN Xiao-feng, GU Zheng-biao . Use of two-stage dough mixing process in improving water distribution of dough and qualities of bread made from wheat–potato flour[J]. >Journal of Integrative Agriculture, 2021, 20(1): 300-310.
[15] WANG Yi-fan, LIAO Yu-qiu, WANG Ya-peng, YANG Jiang-wei, ZHANG Ning, SI Huai-jun. Genome-wide identification and expression analysis of StPP2C gene family in response to multiple stresses in potato (Solanum tuberosum L.)[J]. >Journal of Integrative Agriculture, 2020, 19(6): 1609-1624.
No Suggested Reading articles found!