Nitric Oxide Mediates Brassinosteroids-induced Chilling Tolerance in Chorispora bungeana Suspension Cultured Cells

doi:10.7525/j.issn.1673-5102.2024.01.014

Abstract

Abstract:

In order to investigate whether brassinosteroids（BRs）-induced chilling tolerance is regulated through nitric oxide（NO） signaling molecule， the suspension cultured cells of Chorispora bungeana were treated with 24-epibrassinolide（EBR）， NO donor SNP， NO scavenger PTIO， nitric oxide synthase（NOS） inhibitor L-NAME， EBR+PTIO and EBR+L-NAME respectively， and the effects of the above treatments on chilling tolerance， reactive oxygen species（ROS） levels and antioxidant defense system were analyzed in the cells under low temperature stress. The results showed that：（1）exogenous EBR treatment enhanced chilling tolerance in the suspension cultures and alleviated the inhibition of cell viability and aggravations of ion leakage and membrane lipid peroxidation induced by low temperature. The effects of SNP treatment on the above physiological measures were similar to those of EBR. （2）Application of PTIO or L-NAME in combination with EBR significantly decreased cell viability and increased ion leakage and membrane lipid peroxidation in C. bungeana suspension cultures under chilling stress compared with those of EBR treatment alone， suggesting that the block in NO signaling decreased the EBR-enhanced chilling tolerance. （3）EBR treatment further increased NO production and NOS activity in the suspension cells compared with those under chilling stress alone， whereas the EBR-induced NO signal was quenched by the addition of PTIO or L-NAME. （4）Both EBR and SNP obviously inhibited the increases in hydrogen peroxide（H₂O₂） content， superoxide radical（ $O 2 · –$ ） production rate and hydroxyl radical （OH^-）^{content caused by chilling， and remarkably enhanced the activities of ascorbate peroxidase（APX）， catalase（CAT）， glutathione reductase（GR）， peroxidase（POD） and superoxide dismutase（SOD） and contents of ascorbate （AsA） and glutathione（GSH） in the suspension cultured cells， thus alleviating oxidative injury caused by low temperature. However， PTIO and L-NAME blocked the protective effects of EBR. In conclusion， these results suggested that EBR-induced chilling tolerance in C. bungeana suspension cultured cells was through the promotion of NO accumulation by activating NOS activity. EBR might confer an increased tolerance to chilling stress by suppressing the accumulation of ROS caused by chilling and enhancing antioxidant defense system in the suspension cells， both of which were partially regulated by NO signal， resulting in the alleviation of chilling-induced oxidative damage and membrane lipid peroxidation. Thus， NOS-derived NO might be a downstream signaling molecule of EBR signal in C. bungeana suspension cultured cells under low temperature stress.}

Key words: brassinosteroids, nitric oxide, Chorispora bungeana, chilling tolerance, reactive oxygen species, antioxidant defense system

CLC Number:

Q945.78

Yajie LIU, Lizhe AN. Nitric Oxide Mediates Brassinosteroids-induced Chilling Tolerance in Chorispora bungeana Suspension Cultured Cells[J]. Bulletin of Botanical Research, 2024, 44(1): 118-131.

Figures/Tables 7

Fig.1

Fig.2

Fig.3

Fig.4

Fig.5

Fig.6

Fig.7

References 47

1	项洪涛，郑殿峰，何宁，等.植物对低温胁迫的生理响应及外源脱落酸缓解胁迫效应的研究进展［J］.草业学报，2021，30（1）：208-219.
	XIANG H T， ZHENG D F， HE N，et al.Research progress on the physiological response of plants to low temperature and the amelioration effcectiveness of exogenous ABA［J］.Acta Prataculturae Sinica，2021，30（1）：208-219.
2	SADURA I， JANECZKO A.Physiological and molecular mechanisms of brassinosteroid-induced tolerance to high and low temperature in plants［J］.Biologia Plantarum，2018，62（4）：601-616.
3	XU M J， DONG J F， ZHANG M，et al.Cold-induced endogenous nitric oxide generation plays a role in chilling tolerance of loquat fruit during postharvest storage［J］.Postharvest Biology and Technology，2012，65：5-12.
4	牟雪姣，刘理想，孟鹏鹏，等.外源NO缓解蝴蝶兰低温胁迫伤害的生理机制研究［J］.西北植物学报，2015，35（5）：978-984.
	MU X J， LIU L X， MENG P P，et al.Physiological mechanism of exogenous nitric oxide on alleviating low temperature stress of Phalaenopsis spp.［J］.Acta Botanica Boreali-Occidentalia Sinica，2015，35（5）：978-984.
5	ANWAR A， LIU Y M， DONG R R，et al.The physiological and molecular mechanism of brassinosteroid in response to stress：a review［J］.Biological Research，2018，51（1）：46.
6	黄玉辉，黄如葵，陈小凤，等.油菜素内酯对苦瓜抗冷性生理指标的影响［J］.南方农业学报，2012，43（5）：592-596.
	HUANG Y H， HUANG R K， CHEN X F，et al.Effects of brassinolide on chilling-tolerance in bitter gourd seedlings［J］.Journal of Southern Agriculture，2012，43（5）：592-596.
7	孙玉珺，吴玥，马德志，等.外源油菜素内酯对低温胁迫下玉米发芽及幼苗生理特性的影响［J］.华北农学报，2019，34（3）：119-128.
	SUN Y J， WU Y， MA D Z，et al.Effects of brassinolide on seed germination and seedling physiology of maize under low temperature stress［J］.Acta Agriculturae Boreali-Sinica，2019，34（3）：119-128.
8	黄斌，李文科，李梦露，等.24-表油菜素内酯对低温下黄瓜种子萌发和幼苗低温抗性的影响［J］.中国蔬菜，2021（12）：59-66.
	HUANG B， LI W K， LI M L，et al.Effects of 24-epibrassinolide on seed germination under low temperature and cold stress tolerance of cucumber seedling［J］.China Vegetables，2021（12）：59-66.
9	惠竹梅，王智真，胡勇，等.24-表油菜素内酯对低温胁迫下葡萄幼苗抗氧化系统及渗透调节物质的影响［J］.中国农业科学，2013，46（5）：1005-1013.
	HUI Z M， WANG Z Z， HU Y，et al.Effects of 24-epibrassinolide on the antioxidant system and osmotic adjustment substance in grape seedlings（V.vinifera L.） under chilling stress［J］.Scientia Agricultura Sinica，2013，46（5）：1005-1013.
10	JANECZKO A， HURA K， SKOCZOWSKI A，et al.Temperature-dependent impact of 24-epibrassinolide on the fatty acid composition and sugar content in winter oilseed rape callus［J］.Acta Physiologiae Plantarum，2009，31：71-79.
11	刘亚洁，安黎哲.低温胁迫下油菜素内酯对高山离子芥悬浮细胞膜系统的影响［J］.四川大学学报（自然科学版），2020，57（4）：797-803.
	LIU Y J， AN L Z.Effect of brassinosteroids on the membrane system of Chorispora bungeana suspension-cultured cells under chilling stress［J］.Journal of Sichuan University （Natural Science Edition），2020，57（4）：797-803.
12	张盛敏，王罗霞，陈银华.一氧化氮与植物激素之间相互作用研究进展［J］.南方农业学报，2011，42（3）：261-266.
	ZHANG S M， WANG L X， CHEN Y H.Research progress on the interaction between nitric oxide and phytohormones［J］.Journal of Southern Agriculture，2011，42（3）：261-266.
13	张艳艳，章文华，薛丽，等.一氧化氮在植物生长发育和抗逆过程中的作用研究进展［J］.西北植物学报，2012，32（4）：835-842.
	ZHANG Y Y， ZHANG W H， XUE L，et al.Advance on NO function in plant growth，development and abiotic stress tolerance［J］.Acta Botanica Boreali-Occidentalia Sinica，2012，32（4）：835-842.
14	SAMI F， FAIZAN M， FARAZ A，et al.Nitric oxide-mediated integrative alterations in plant metabolism to confer abiotic stress tolerance，NO crosstalk with phytohormones and NO-mediated post translational modifications in modulating diverse plant stress［J］.Nitric Oxide，2018，73：22-38.
15	张喆慧，王昕，金可默，等.一氧化氮在植物发育及植物-微生物互作中的作用机制研究进展［J］.植物营养与肥料学报，2021，27（4）：706-718.
	ZHANG Z H， WANG X， JIN K M，et al.Advances in studies on the role of nitric oxide in plant development and plant-microbial interactions［J］.Journal of Plant Nutrition and Fertilizers，2021，27（4）：706-718.
16	陈银萍，王晓梅，杨宗娟，等.NO对低温胁迫下玉米种子萌发及幼苗生理特性的影响［J］.农业环境科学学报，2012，31（2）：270-277.
	CHEN Y P， WANG X M， YANG Z J，et al.Effects of nitric oxide on seed germination and physiological reaction of maize seedlings under low temperature stress［J］.Journal of Agro-Environment Science，2012，31（2）：270-277.
17	何文平，王新霞，张旭强，等.低温胁迫下外源脱落酸对高山离子芥抗氧化酶和渗透调节物质的影响［J］.植物研究，2016，36（6）：870-877.
	HE W P， WANG X X， ZHANG X Q，et al.Exogenous ABA affected on antioxidant enzymes and osmotic adjustment in Chorispora bungeana under low temperature stress［J］.Bulletin of Botanical Research，2016，36（6）：870-877.
18	GUO F X， ZHANG M X， CHEN Y，et al.Relation of several antioxidant enzymes to rapid freezing resistance in suspension cultured cells from alpine Chorispora bungeana ［J］.Cryobiology，2006，52（2）：241-250.
19	ZHAO L， HE J X， WANG X M，et al.Nitric oxide protects against polyethylene glycol-induced oxidative damage in two ecotypes of reed suspension cultures［J］.Journal of Plant Physiology，2008，165（2）：182-191.
20	李合生.植物生理生化实验原理和技术［M］.北京：高等教育出版社，2000：260-261.
	LI H S.Principles and techniques of plant physiological and biochemical experiments［M］.Beijing：Higher Education Press，2000：260-261.
21	LIU Y J， JIANG H F， ZHAO Z G，et al.Nitric oxide synthase like activity-dependent nitric oxide production protects against chilling-induced oxidative damage in Chorispora bungeana suspension cultured cells［J］.Plant Physiology and Biochemistry，2010，48（12）：936-944.
22	LI J， YANG P， GAN Y T，et al.Brassinosteroid alleviates chilling-induced oxidative stress in pepper by enhancing antioxidation systems and maintenance of photosystem II［J］.Acta Physiologiae Plantarum，2015，37：222.
23	杨萍，李杰.2，4-表油菜素内酯对低温胁迫下辣椒幼苗抗氧化系统的影响［J］.北方园艺，2017（21）：7-12.
	YANG P， LI J.Effect of 2，4-epibrassinolide on antioxidant system of pepper seedlings under low temperature stress［J］.Northern Horticulture，2017（21）：7-12.
24	普布卓玛，罗艺岚，高金柱，等.2，4表-油菜素内酯对低温胁迫下西藏野生垂穗披碱草幼苗抗氧化保护和渗透调节的影响［J］.草地学报，2019，27（3）：547-552.
	PUBU Z M， LUO Y L， GAO J Z，et al.Effect of 2，4-epibrassinolide on antioxidant defense and osmotic adjustment of Elymus nutans under low temperature stress［J］.Acta Agrestia Sinica，2019，27（3）：547-552.
25	ALI B， HAYAT S， FARIDUDDIN Q，et al.2，4-epibrassinolide protects against the stress generated by salinity and nickel in Brassica juncea ［J］.Chemosphere，2008，72（9）：1387-1392.
26	SUN B T， JING Y， CHEN K M，et al.Protective effect of nitric oxide on iron deficiency-induced oxidative stress in maize（Zea mays）［J］.Journal of Plant Physiology，2007，164（5）：536-543.
27	张政委，索琳格，吴佩，等.Ca²⁺参与外源NO增强低温胁迫下黄瓜幼苗叶片抗氧化能力［J］.核农学报，2018，32（3）：600-608.
	ZHANG Z W， SUO L G， WU P，et al.Ca²⁺ involved in antioxidant enzymes activities of cucumber seedling leaves enhanced by exogenous nitric oxide under low temperature stress［J］.Journal of Nuclear Agricultural Sciences，2018，32（3）：600-608.
28	黄志明，陈宇，吴晶晶，等.硝普钠对低温胁迫下枇杷幼果线粒体AsA-GSH循环代谢的影响［J］.热带作物学报，2011，32（8）：1469-1474.
	HUANG Z M， CHEN Y， WU J J，et al.Effects of exogenous sodium nitroprusside on AsA-GSH circulation metabolism in mitochondria of young loquat fruits under low temperature stress［J］.Chinese Journal of Tropical Crops，2011，32（8）：1469-1474.
29	陈宇，林素英，黄志明，等.枇杷幼果内源一氧化氮和茉莉酸对低温胁迫的响应［J］.植物科学学报，2012，30（6）：611-617.
	CHEN Y， LIN S Y， HUANG Z M，et al.Response of endogenous nitric oxide and jasmonate acid to low temperature stress in young loquat fruits［J］.Plant Science Journal，2012，30（6）：611-617.
30	LOMBARDO M C， LAMATTINA L.Abscisic acid and nitric oxide modulate cytoskeleton organization，root hair growth and ectopic hair formation in Arabidopsis ［J］.Nitric Oxide，2018，80：89-97.
31	PAGNUSSAT G C， SIMONTACCHI M， PUNTARULO S，et al.Nitric oxide is required for root organogenesis［J］.Plant Physiology，2002，129（3）：954-956.
32	CARIMI F， ZOTTINI M， COSTA A，et al.NO signalling in cytokinin-induced programmed cell death［J］.Plant，Cell and Environment，2005，28（9）：1171-1178.
33	刘国华，刘菁，侯丽霞，等.NO可能作为Ca²⁺的下游信号介导乙烯诱导的蚕豆气孔关闭［J］.分子细胞生物学报，2009，42（2）：145-155.
	LIU G H， LIU J， HOU L X，et al.NO may function in the downstream of Ca²⁺ in ethylene induced stomatal closure in Vicia faba L.［J］.Journal of Molecular Cell Biology，2009，42（2）：145-155.
34	ASGHER M， PER T S， MASOOD A，et al.Nitric oxide signaling and its crosstalk with other plant growth regulators in plant responses to abiotic stress［J］.Environmental Science and Pollution Research，2017，24：2273-2285.
35	SINGHAL R K， JATAV H S， AFTAB T，et al.Roles of nitric oxide in conferring multiple abiotic stress tolerance in plants and crosstalk with other plant growth regulators［J］.Journal of Plant Growth Regulation，2021，40：2303-2328.
36	TOSSI V， LAMATTINA L， CASSIA R.Pharmacological and genetical evidence supporting nitric oxide requirement for 2，4-epibrassinolide regulation of root architecture in Arabidopsis thaliana ［J］.Plant Signaling and Behavior，2013，8（7）：e24712.
37	LI X， ZHANG L， AHAMMED G J，et al.Nitric oxide mediates brassinosteroid-induced flavonoid biosynthesis in Camellia sinensis L.［J］.Journal of Plant Physiology，2017，214：145-151.
38	李雨桐，廖伟彪，胡琳莉，等.一氧化氮（NO）在油菜素内酯（BR）诱导黄瓜不定根发生过程中的作用［J］.甘肃农业大学学报，2018，53（6）：98-104，113.
	LI Y T， LIAO W B， HU L L，et al.Nitric oxide participates in brassinolide inducing adventitious root of cucumber［J］.Journal of Gansu Agricultural University，2018，53（6）：98-104，113.
39	ZHU T， DENG X G， TAN W R，et al.Nitric oxide is involved in brassinosteroid-induced alternative respiratory pathway in Nicotiana benthamiana seedlings’ response to salt stress［J］.Physiologia Plantarum，2016，156（2）：150-163.
40	ZHANG A Y， ZHANG J， ZHANG J H，et al.Nitric oxide mediates brassinosteroid-induced ABA biosynthesis involved in oxidative stress tolerance in maize leaves［J］.Plant and Cell Physiology，2011，52（1）：181-192.
41	KAYA C， ASHRAF M， WIJAYA L，et al.The putative role of endogenous nitric oxide in brassinosteroid-induced antioxidant defence system in pepper （Capsicum annuum L.） plants under water stress［J］.Plant Physiology and Biochemistry，2019，143：119-128.
42	CUI J X， ZHOU Y H， DING J G，et al.Role of nitric oxide in hydrogen peroxide-dependent induction of abiotic stress tolerance by brassinosteroids in cucumber［J］.Plant，Cell and Environment，2011，34（2）：347-358.
43	KAYA C， ASHRAF M， ALYEMENI M N，et al.The role of nitrate reductase in brassinosteroid-induced endogenous nitric oxide generation to improve cadmium stress tolerance of pepper plants by upregulating the ascorbate-glutathione cycle［J］.Ecotoxicology and Environmental Safety，2020，196：110483.
44	马金虎，邢国芳，杨小环，等.外源EBR和NO信号对低温胁迫下玉米种胚抗氧化系统和低温响应基因表达的影响［J］.应用生态学报，2015，26（5）：1411-1418.
	MA J H， XING G F， YANG X H，et al.Effects of exogenous EBR and NO signal on antioxidant system and low response gene expression under cold stress on maize embryo［J］.Chinese Journal of Applied Ecology，2015，26（5）：1411-1418.
45	REDDY A S N， ALI G S， CELESNIK H，et al.Coping with stresses：roles of calcium- and calcium/calmodulin-regulated gene expression［J］.The Plant Cell，2011，23（6）：2010-2032.
46	柯媛媛，陈翔，倪芊芊，等.低温逆境胁迫下小麦ROS代谢及调控机制研究进展［J］.大麦与谷类科学，2021，38（1）：1-6，21.
	KE Y Y， CHEN X， NI Q Q，et al.Research progress of the metabolism of reactive oxygen species and its regulation mechanisms in wheat under low temperature stress［J］.Barley and Cereal Sciences，2021，38（1）：1-6，21.
47	AN S M， LIU Y， SANG K Q，et al.Brassinosteroid signaling positively regulates abscisic acid biosynthesis in response to chilling stress in tomato［J］.Journal of Integrative Plant Biology，2023，65（1）：10-24.

[1]	Hong YANG, Lifeng WANG, Longjun DAI, Bingbing GUO. Effects of Tapping Panel Dryness on Mitochondrial Ultrastructure and ROS Metabolism in Barks of Rubber Tree (Hevea brasiliensis) [J]. Bulletin of Botanical Research, 2023, 43(1): 69-75.
[2]	Jianxin LIU, Ruirui LIU, Xiuli LIU, Haiyan JIA, Ting BU, Na LI. Effects of Spraying NaHS at Different Growth Stages on H₂S Production and Reactive Oxygen Species Metabolism of Naked Oat Leaves under Saline-Alkali Stress [J]. Bulletin of Botanical Research, 2022, 42(3): 455-465.
[3]	Xiao-Ju ZHAO, Yi-Ting ZHANG, Jia LIU, Yang LIU, Zhong-Hua TANG. Regulation of NO in the Phenolics Metabolism of Catharanthus roseus under Salt Stress [J]. Bulletin of Botanical Research, 2021, 41(4): 633-640.
[4]	ZHAO Min, ZHAO Qi-An, LIU Bo, WANG Yue-Xuan, ZHANG Li-Huan, YANG Ning. Effect of H₂S on Chorispora bungeana under Drought Stress [J]. Bulletin of Botanical Research, 2018, 38(6): 902-912.
[5]	ZHAO Xiao-Ju, ZHANG Li-Xia, MAN Xiu-Ling. Effects of Exogenous NO on Seed Germination and Physiological Metabolism in Catharanthus roseus Seedling under NaCl Stress [J]. Bulletin of Botanical Research, 2018, 38(5): 669-674.
[6]	LIU Jian-Xin, OU Xiao-Bin, LIU Xiu-Li, WANG Jin-Cheng. Principal Component and Subordinate Function of the Alleviating Effects of Hydrogen Peroxide(H₂O₂) on Low-temperature Stress in Naked Oat(Avena nuda) Seedlings [J]. Bulletin of Botanical Research, 2018, 38(5): 748-756.
[7]	ZHANG Xu-Qiang, ZHAO Min, CHEN Lu, AN Yan-Huang, YANG Peng-Jun, Wang Xin-Xia, YANG Ning. Relationship Between Phospholipase Dδ and Nitric Oxide Responding to the Seed Germination in Arabidopsis thaliana under Drought Stress [J]. Bulletin of Botanical Research, 2018, 38(4): 526-534.
[8]	ZHANG Li-Li, ZHANG Fu-Chun. Transcriptomic Analysis of the Halostachys caspica in Response to Short-term Salt Stress [J]. Bulletin of Botanical Research, 2018, 38(1): 91-99.
[9]	LI Mei-Lan;LI De-Wen;YU Jing-Hua;ZU Yuan-Gang. Effect of Exogenous Nitric Oxide on Photosynthetic Pigment and Antioxidant Enzymes in Taxus chinensis* var. marei Seedlings [J]. Bulletin of Botanical Research, 2013, 33(1): 39-44.
[10]	LIU Jian-Xin;WANG Rui-Juan;JIA Hai-Yan;. Alleviation of SNP on Growth Inhibition and Root Oxidative Damage Caused by Lanthanum in Ryegrass Seedlings [J]. Bulletin of Botanical Research, 2012, 32(6): 680-684.
[11]	LIU Jian-Xin;WANG Xin;WANG Rui-Juan;LI Dong-Bo. Effects of Alkaline Stress on the Metabolism of Reactive Oxygen Species and Osmotica Accumulation in Ryegrass Seedling Roots [J]. Bulletin of Botanical Research, 2011, 31(6): 674-679.
[12]	LIU Jian-Xin;WANG Xin;LI Bo-Ping. Effects of Exogenous Nitric Oxide Donor on the Ascorbate-glutathione Cycle in Peganum multisectum Seedling Leaves under NaCl Stress [J]. Bulletin of Botanical Research, 2010, 30(1): 37-41.
[13]	YANG Ying-Li;FAN Qing;WEI Xue-Ling;SHI Ru-Xia;MA Xu-Jun;ZHANG Sai-Na. Effect of Sodium Nitroprusside on Lipid Peroxidation and Antioxidant Enzyme Activity of Nitraria tangutorum Bobr. Callus [J]. Bulletin of Botanical Research, 2009, 29(3): 303-307.
[14]	LIU Jian-Xin;HU Hao-Bin;WANG Xin. Effects of Exogenous Nitric Oxide on Active Oxygen Metabolism, Polyamine Content and Photosynthesis of Ryegrass(Lolium perenne L.) Seedlings Under Salt Stress [J]. Bulletin of Botanical Research, 2009, 29(3): 313-319.
[15]	LI Cai-Xiang;HUI Wei*;TU Yin-Hua;NIU Rui-Xue;JIN Hong. Effect of Nitric Oxide（NO）Fumigation on the Photosynthesis of Garlic Sprouts [J]. Bulletin of Botanical Research, 2009, 29(3): 308-312.