<i>GhLhcb2A1</i>基因在陆地棉低温和干旱响应中的功能

doi:10.7668/hbnxb.20195341

摘要/Abstract

摘要：

捕光叶绿素a/b结合蛋白在植物光合进程及非生物胁迫响应中发挥着重要的作用。为了研究陆地棉GhLhcb2A1基因特征、表达特性以及在低温和干旱响应中的功能,以冀棉262叶片cDNA为模板进行PCR扩增,获得了GhLhcb2A1基因的CDS全长,通过生物信息学分析了基因及其编码蛋白的基本特征,利用qRT-PCR技术检测了基因的组织表达特性以及低温和干旱响应表达模式,采用病毒诱导的基因沉默技术验证了GhLhcb2A1基因在低温和干旱响应中的功能。研究结果表明,GhLhcb2A1基因CDS全长为798 bp,编码265个氨基酸。GhLhcb2A1在叶片中高表达,在低温和干旱处理的叶片和根中显著上调表达,并在低温和干旱处理3 h的叶片中达到最大值,分别是对照叶片中的17.42,30.03倍,在低温处理6 h和干旱处理12 h的根中达到最大值,分别是对照根中的11.65,65.04倍。亚细胞定位结果表明,GhLhcb2A1蛋白在细胞叶绿体中表达。GhLhcb2A1基因沉默植株与对照植株相比,低温和干旱处理造成的植株失水干枯等表型更严重,叶片积累的丙二醛含量显著升高,脯氨酸含量和超氧化物歧化酶活性则显著降低,说明GhLhcb2A1基因沉默植株对低温和干旱胁迫的抵抗力降低。以上研究表明,GhLhcb2A1基因在低温和干旱响应中发挥正调控的作用。

关键词: 陆地棉, GhLhcb2A1, 低温, 干旱, 捕光叶绿素a/b结合蛋白

Abstract:

Light-harvesting chlorophyll a/b binding proteins are important in plant photosynthesis and abiotic stress response.To study the characteristics of GhLhcb2A1 and its expression patterns and functions in low temperature and drought response in upland cotton,full-length CDS of GhLhcb2A1 gene was cloned from the leaf cDNA of Jimian 262 by PCR.Bioinformatic analysis was conducted to learn the basic characteristics of the gene.The expression patterns and functions in low temperature and drought response were evaluated by qRT-PCR and virus-induced gene silencing.It was shown that the length of GhLhcb2A1 CDS was 798 bp,encoding 265 amino acids.GhLhcb2A1 was highly expressed in leaves and was significantly up-regulated in leaves and roots under low temperature and drought treatment.Compared with the control,its expression maximized at 3 h under low temperature and drought in leaves with 17.42 and 30.03 folds increase respectively,whereas maximized at 6 h under low temperature and 12 h under drought in roots with 11.65 and 65.04 folds respectively.Subcellular localization assay verified that GhLhcb2A1 was expressed in the chloroplasts of cells.Compared with the control plants,GhLhcb2A1 silenced plants showed a more severe phenotype of water loss and dryness under both low temperatures and drought.The accumulated malondialdehyde content in the leaves of the silenced plants was significantly higher than that of the control,while the proline content and superoxide dismutase activity were significantly lower than those of the controls,suggesting that GhLhcb2A1 silenced plant reduced the resistance to low temperature and drought.The above results implied that this gene played a positive role in regulating low temperature and drought response.

Key words: Upland cotton, GhLhcb2A1, Low temperature, Drought, Light-harvesting chlorophyll a/b binding protein

蔡肖, 刘存敬, 张素君, 李兴河, 王海涛, 唐丽媛, 张建宏. GhLhcb2A1基因在陆地棉低温和干旱响应中的功能[J]. 华北农学报, 2025, 40(1): 14-21. doi: 10.7668/hbnxb.20195341.

CAI Xiao, LIU Cunjing, ZHANG Sujun, LI Xinghe, WANG Haitao, TANG Liyuan, ZHANG Jianhong. Function Analysis of GhLhcb2A1 Gene in Response to Low Temperature and Drought in Upland Cotton[J]. Acta Agriculturae Boreali-Sinica, 2025, 40(1): 14-21. doi: 10.7668/hbnxb.20195341.

http://www.hbnxb.net/CN/Y2025/V40/I1/14

图/表 6

表1 引物序列

Tab.1 Primer sequences

引物名称 Prime name	序列(5'—3') Sequence (5'—3')	用途 Function
GhLhcb2A1-F	ATGGCAACCTCTGCTATCCA	基因克隆
GhLhcb2A1-R	TCATTTTCCGGGGACAAAGT
GhLhcb2A1-q-F	GTTGGCCAGACCGCTTTGAA	实时荧光定量PCR
GhLhcb2A1-q-R	CATGGTAACACGACCACGATTG
GhLhcb2A1-GFP-F	TTCCCGGGATGGCAACCTCTGCTATCCA	GFP融合载体构建
GhLhcb2A1-GFP-R	TTGGATCCTTTTCCGGGGACAAAGTTAG
GhUBQ7-F	GAAGGCATTCCACCTGACCAAC	实时荧光定量PCR
GhUBQ7-R	CTTGACCTTCTTCTTCTTGTGCTTG

图1 GhLhcb2A1蛋白磷酸化位点预测和跨膜区预测

Fig.1 Prediction of phosphorylation sites and transmembrane regions in GhLhcb2A1 protein

图2 GhLhcb2A1蛋白三级结构

Fig.2 Tertiary structure of GhLhcb2A1 protein

图3 GhLhcb2A1基因组织表达特性及低温、干旱响应表达分析采用最小显著差法(LSD)检验,不同小写字母代表不同样本间存在显著差异(P<0.05)。图5同。

Fig.3 Expression of GhLhcb2A1 gene in different tissues,low temperature and drought response The least significant difference(LSD)test was used,lowercase letters represent significant differences between samples(P<0.05).The same as Fig.5.

图4 GhLhcb2A1亚细胞定位

Fig.4 Subcellular localization of GhLhcb2A1 protein

图5 GhLhcb2A1基因沉默植株对低温和干旱胁迫的响应 A.阳性对照GhCLA1基因沉默植株叶片漂白表型;B.qRT-PCR检测VIGS接种植株叶片GhLhcb2A1基因表达量;C.GhLhcb2A1基因沉默和阴性对照植株胁迫处理0 h表型;D.GhLhcb2A1基因沉默和阴性对照植株低温胁迫处理 7 d表型;E.GhLhcb2A1基因沉默和阴性对照植株干旱处理10 d表型。

Fig.5 Response of GhLhcb2A1 silencing plants to low temperature and drought A.Positive control of GhCLA1 silencing plants showed green loss in the leaf;B.Expression analysis of GhLhcb2A1 gene in VIGS plant leaves by qRT-PCR;C.Phenotype of GhLhcb2A1 silencing plants and negative control plants under stress at 0 h;D.Phenotype of GhLhcb2A1 silencing plants and negative control plants after 7 d under cold stress;E.Phenotype of GhLhcb2A1 silencing plants and negative control plants after 10 d under drought stress.

参考文献 32

[1]	Lan Y H, Song Y, Zhao F, Cao Y, Luo D N, Qiao D R, Cao Y, Xu H. Phylogenetic,structural and functional evolution of the LHC gene family in plant species[J]. International Journal of Molecular Sciences, 2023, 24(1):488.doi:10.3390/ijms24010488.
[2]	Jansson S. The light-harvesting chlorophyll ab-binding proteins[J]. Biochimica et Biophysica Acta, 1994, 1184(1):1-19.doi:10.1016/0005-2728(94)90148-1. pmid: 8305447
[3]	Jansson S. A guide to the Lhc genes and their relatives in Arabidopsis[J]. Trends in Plant Science, 1999, 4(6):236-240.doi:10.1016/s1360-1385(99)01419-3.
[4]	Klimmek F, Sjödin A, Noutsos C, Leister D, Jansson S. Abundantly and rarely expressed Lhc protein genes exhibit distinct regulation patterns in plants[J]. Plant Physiology, 2006, 140(3):793-804.doi:10.1104/pp.105.073304. pmid: 16524980
[5]	裴雅婷, 刘玉霖, 李世伟, 张瑀瑶, 高红秀, 唐鑫华, 石瑛. 表油菜素内酯对遮光处理下马铃薯生理、产量及LHcb基因表达的影响[J]. 华北农学报, 2024, 39(4):133-142.doi:10.7668/hbnxb.20194796.
	Pei Y T, Liu Y L, Li S W, Zhang Y Y, Gao H X, Tang X H, Shi Y. Effects of epbrassinolide on physiology,yield and LHcb gene expression of potato under shading treatment[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(4):133-142. doi: 10.7668/hbnxb.20194796
[6]	Pietrzykowska M, Suorsa M, Semchonok D A, Tikkanen M, Boekema E J, Aro E M, Jansson S. The light-harvesting chlorophyll a/b binding proteins Lhcb1 and Lhcb2 play complementary roles during state transitions in Arabidopsis[J]. The Plant Cell, 2014, 26(9):3646-3660.doi:10.1105/tpc.114.127373. pmid: 25194026
[7]	Rochaix J D. Regulation and dynamics of the light-harvesting system[J]. Annual Review of Plant Biology, 2014, 65:287-309.doi:10.1146/annurev-arplant-050213-040226.
[8]	赵奇, 茹京娜, 李宜统, 王超, 徐兆师, 王睿辉. 小麦Lhc基因家族鉴定与表达模式分析[J]. 植物遗传资源学报, 2022, 23(6):1766-1781.doi:10.13430/j.cnki.jpgr.20220401001.
	Zhao Q, Ru J N, Li Y T, Wang C, Xu Z S, Wang R H. Identification and expression pattern analysis of lhc gene family members in wheat[J]. Journal of Plant Genetic Resources, 2022, 23(6):1766-1781.
[9]	Chen L P, Yang W B, Liu S Q, Meng Y, Zhu Z H, Liang R, Cao K Y, Xie Y Z, Li X J. Genome-wide analysis and identification of light-harvesting chlorophyll a/b binding(LHC)gene family and BSMV-VIGS silencing TaLHC86 reduced salt tolerance in wheat[J]. International Journal of Biological Macromolecules, 2023, 242:124930.doi:10.1016/j.ijbiomac.2023.124930.
[10]	Hu Z H, Zhang N, Qin Z Y, Li J W, Tao J P, Yang N, Chen Y, Kong J Y, Luo W, Chen X, Li X H, Xiong A S, Zhuang J. Circadian rhythm response and its effect on photosynthetic characteristics of the Lhcb family genes in tea plant[J]. BMC Plant Biology, 2024, 24(1):333.doi:10.1186/s12870-024-04958-0.
[11]	Li X L, Jiang Z Y, Zhang C F, Cai K F, Wang H, Pan W Y, Sun X P, Gao Y B, Xu K. Comparative genomics analysis provides insights into evolution and stress responses of Lhcb genes in Rosaceae fruit crops[J]. BMC Plant Biology, 2023, 23(1):484.doi:10.1186/s12870-023-04438-x.
[12]	Luo J, Abid M, Tu J, Gao P X, Wang Z P, Huang H W. Genome-wide identification of the LHC gene family in kiwifruit and regulatory role of AcLhcb3.1/3.2 for chlorophyll a content[J]. International Journal of Molecular Sciences, 2022, 23(12):6528.doi:10.3390/ijms23126528.
[13]	Umate P. Genome-wide analysis of the family of light-harvesting chlorophyll a/b-binding proteins in Arabidopsis and rice[J]. Plant Signaling & Behavior, 2010, 5(12):1537-1542.doi:10.4161/psb.5.12.13410.
[14]	Xu Y H, Liu R, Yan L, Liu Z Q, Jiang S C, Shen Y Y, Wang X F, Zhang D P. Light-harvesting chlorophyll a/b-binding proteins are required for stomatal response to abscisic acid in Arabidopsis[J]. Journal of Experimental Botany, 2012, 63(3):1095-1106.doi:10.1093/jxb/err315. pmid: 22143917
[15]	Zhao S, Gao H B, Luo J W, Wang H B, Dong Q L, Wang Y P, Yang K Y, Mao K, Ma F W. Genome-wide analysis of the light-harvesting chlorophyll a/b-binding gene family in apple(Malus domestica)and functional characterization of MdLhcb4.3,which confers tolerance to drought and osmotic stress[J]. Plant Physiology and Biochemistry, 2020, 154:517-529.doi:10.1016/j.plaphy.2020.06.022.
[16]	Wang L, Wei J, Shi X Y, Qian W H, Mehmood J, Yin Y M, Jia H J. Identification of the light-harvesting chlorophyll a/b binding protein gene family in peach(Prunus persica L.)and their expression under drought stress[J]. Genes, 2023, 14(7):1475.doi:10.3390/genes14071475.
[17]	Ye J J, Lin X Y, Yang Z X, Wang Y Q, Liang Y R, Wang K R, Lu J L, Lu P, Zheng X Q. The light-harvesting chlorophyll a/b-binding proteins of photosystem Ⅱ family members are responsible for temperature sensitivity and leaf color phenotype in albino tea plant[J]. Journal of Advanced Research, 2024, 66:87-104.doi:10.1016/j.jare.2023.12.017.
[18]	Deng Y S, Kong F Y, Zhou B, Zhang S, Yue M M, Meng Q W. Heterology expression of the tomato LeLhcb2 gene confers elevated tolerance to chilling stress in transgenic tobacco[J]. Plant Physiology and Biochemistry, 2014, 80:318-327.doi:10.1016/j.plaphy.2014.04.017.
[19]	刘亚丽, 张凯敏, 高彩球. 刚毛柽柳光捕获叶绿素a/b结合蛋白基因Thcab1的克隆与分析[J]. 东北林业大学学报, 2014, 42(2):52-56.doi:10.13759/j.cnki.dlxb.2014.02.013.
	Liu Y L, Zhang K M, Gao C Q. Chlorophyll a/b binding proteins Thcab1 gene clone and expression analysis of Tamarix hispida[J]. Journal of Northeast Forestry University, 2014, 42(2):52-56.
[20]	翟玉山, 邓宇晴, 董萌, 徐倩, 程光远, 彭磊, 林彦铨, 徐景升. 甘蔗捕光叶绿素a/b结合蛋白基因ScLhca3的克隆及表达[J]. 作物学报, 2016, 42(9):1332-1341.doi:10.3724/SP.J.1006.2016.01332.
	Zhai Y S, Deng Y Q, Dong M, Xu Q, Cheng G Y, Peng L, Lin Y Q, Xu J S. Cloning and characterization of light harvesting chlorophylla/b-binding protein coding gene(ScLhca3)in sugarcane[J]. Acta Agronomica Sinica, 2016, 42(9):1332-1341.
[21]	Liu M X, Zhang S B, Hu J X, Sun W X, Padilla J, He Y L, Li Y, Yin Z Y, Liu X Y, Wang W H, Shen D Y, Li D Y, Zhang H F, Zheng X B, Cui Z L, Wang G L, Wang P, Zhou B, Zhang Z G. Phosphorylation-guarded light-harvesting complex Ⅱ contributes to broad-spectrum blast resistance in rice[J]. Proceedings of the National Academy of Sciences of the United States of America, 2019, 116(35):17572-17577.doi:10.1073/pnas.1905123116.
[22]	Mehari T G, Xu Y C, Magwanga R O, Umer M J, Kirungu J N, Cai X Y, Hou Y Q, Wang Y H, Yu S X, Wang K B, Zhou Z L, Liu F. Genome wide identification and characterization of light-harvesting chloro a/b binding(LHC)genes reveals their potential role in enhancing drought tolerance in Gossypium hirsutum[J]. Journal of Cotton Research, 2021, 4(1):15.doi:10.1186/s42397-021-00090-8.
[23]	Zhang Q, Ma C, Wang X, Ma Q, Fan S, Zhang C. Genome-wide identification of the light-harvesting chlorophyll a/b binding(Lhc)family in Gossypium hirsutum reveals the influence of GhLhcb2.3 on chlorophyll a synthesis[J]. Plant Biology, 2021, 23(5):831-842.doi:10.1111/plb.13294. pmid: 34263979
[24]	Zhang T Z, Hu Y, Jiang W K, Fang L, Guan X Y, Chen J D, Zhang J B, Saski C A, Scheffler B E, Stelly D M, Hulse-Kemp A M, Wan Q, Liu B L, Liu C X, Wang S, Pan M Q, Wang Y K, Wang D W, Ye W X, Chang L J, Zhang W P, Song Q X, Kirkbride R C, Chen X Y, Dennis E, Llewellyn D J, Peterson D G, Thaxton P, Jones D C, Wang Q, Xu X Y, Zhang H, Wu H T, Zhou L, Mei G F, Chen S Q, Tian Y, Xiang D, Li X H, Ding J, Zuo Q Y, Tao L N, Liu Y C, Li J, Lin Y, Hui Y Y, Cao Z S, Cai C P, Zhu X F, Jiang Z, Zhou B L, Guo W Z, Li R Q, Chen Z J. Sequencing of allotetraploid cotton(Gossypium hirsutum L.acc.TM-1)provides a resource for fiber improvement[J]. Nature Biotechnology, 2015, 33(5):531-537.doi:10.1038/nbt.3207.
[25]	Mittler R, Zandalinas S I, Fichman Y, van Breusegem F. Reactive oxygen species signalling in plant stress responses[J]. Nature Reviews Molecular Cell Biology, 2022, 23(10):663-679.doi:10.1038/s41580-022-00499-2.
[26]	Zhu J K. Abiotic stress signaling and responses in plants[J]. Cell, 2016, 167(2):313-324.doi:10.1016/j.cell.2016.08.029.
[27]	Mignolet-Spruyt L, Xu E J, Idänheimo N, Hoeberichts F A, Mühlenbock P, Brosché M, Van Breusegem F, Kangasjärvi J. Spreading the news:subcellular and organellar reactive oxygen species production and signalling[J]. Journal of Experimental Botany, 2016, 67(13):3831-3844.doi:10.1093/jxb/erw080. pmid: 26976816
[28]	Lei Y, Li B L, Wang X M, Wei J Y, Wang P Y, Zhao J, Yu F, Qi Y F. Chloroplast SRP54 and FtsH protease coordinate thylakoid membrane-associated proteostasis in Arabidopsis[J]. Plant Physiology, 2023, 192(3):2318-2335.doi:10.1093/plphys/kiad199.
[29]	Wang X Y, Wu Z H, Zhou Q, Wang X, Song S, Dong S K. Physiological response of soybean plants to water deficit[J]. Frontiers in Plant Science, 2022, 12:809692.doi:10.3389/fpls.2021.809692.
[30]	Sena F, Monza J, Signorelli S. Determination of free proline in plants[J]. Methods in Molecular Biology, 2024, 2798:183-194.doi:10.1007/978-1-0716-3826-2_12. pmid: 38587743
[31]	Wang Y, Branicky R, Noë A, Hekimi S. Superoxide dismutases:dual roles in controlling ROS damage and regulating ROS signaling[J]. Journal of Cell Biology, 2018, 217(6):1915-1928.doi:10.1083/jcb.201708007.
[32]	Zaib P, Ahmad H M, Attacha S, Rahman M U, Shafiq M R, Parveen K, Fiaz S, Attia K A, Ishaq S, Arif S, Abushady A M, Umer M J. Comparative genomics of light harvesting chlorophyll(LHC)gene family and impact of chlorophyll-a contents under drought stress in Helianthus annuus[J]. Journal of Plant Physiology, 2023, 291:154136.doi:10.1016/j.jplph.2023.154136.

选择文件类型/文献管理软件名称

选择包含的内容

GhLhcb2A1基因在陆地棉低温和干旱响应中的功能

Function Analysis of GhLhcb2A1 Gene in Response to Low Temperature and Drought in Upland Cotton

RichHTML

PDF

可视化

被引次数

摘要/Abstract

引用本文

使用本文

图/表 6

参考文献 32

相关文章 15

编辑推荐

Metrics

本文评价

[1]	赵斌, 姚华, 石娜娜, 高转转, 杨茂, 冯江华, 沈海涛. 甘草4CL基因家族鉴定及其在干旱胁迫下的表达分析[J]. 华北农学报, 2024, 39(S1): 1-10.
[2]	庞志远, 程宇坤, 郭小玲, 任毅, 耿洪伟. 水旱条件下小麦分蘖相关性状全基因组关联分析[J]. 华北农学报, 2024, 39(6): 64-75.
[3]	郭昭阳, 殷宇航, 刘钰, 解一桐, 裴玉贺, 宋希云, 赵美爱. *玉米ZmPAP26b基因的克隆及其在干旱胁迫下表达模式分析*[J]. 华北农学报, 2024, 39(4): 10-19.
[4]	郑鑫鑫, 张艳, 解美霞, 张冬梅, 吴立强, 王省芬, 杨君. *陆地棉抗黄萎病基因GhENODL6互作蛋白筛选*[J]. 华北农学报, 2024, 39(3): 173-178.
[5]	温海洋, 朱紫童, 湛佳伟, 李长, 武博寒, 杨永霞, 张松涛, 贾宏昉. *烟草转录因子NtWRKY11基因克隆、亚细胞定位及表达模式分析*[J]. 华北农学报, 2024, 39(3): 60-66.
[6]	杨朝伟, 孙伟红, 任伟, 王丹, 安明珠, 耿飞龙, 王显国. 抗寒锻炼期和返青期黑麦对低温的生理响应机制[J]. 华北农学报, 2024, 39(3): 104-113.
[7]	黄明, 姜沛沛, 张振旺, 吴金芝, 李友军. 干旱和品种对小麦灌浆期旗叶下午光合速率、关键酶活性和产量的影响[J]. 华北农学报, 2024, 39(2): 90-98.
[8]	张自阳, 周谦, 王依, 王志伟, 朱启迪, 茹振钢, 刘明久. *小麦倒春寒响应基因TaJAZ6的克隆、表达模式及亚细胞定位分析*[J]. 华北农学报, 2024, 39(1): 27-36.
[9]	朱凡, 徐芷琪, 段雨洁, 李阳阳, 宋有洪. 玉米花丝响应干旱胁迫的转录组学分析[J]. 华北农学报, 2024, 39(1): 37-47.
[10]	李晨宇, 足木热木·吐尔逊, 李晓荣, 杨洋, 李波, 于月华. *棉花GhMYB42基因的克隆及原核表达*[J]. 华北农学报, 2024, 39(1): 48-54.
[11]	葛雯冬, 王腾辉, 高美玲, 范震宇, 王玉书. 工业大麻HD-Zip基因家族鉴定与干旱胁迫下的表达分析[J]. 华北农学报, 2024, 39(1): 55-62.
[12]	赵爽, 葛朝红, 石鹤飞, 闵卓, 王广鹏, 李伟明. 板栗WRKY基因家族鉴定及其在干旱胁迫下的表达分析[J]. 华北农学报, 2024, 39(1): 72-82.
[13]	杨宇, 刘雨晨, 周美春, 程璧瑄, 宋建国, 罗乐, 于超. 低温沙藏下密刺蔷薇种子内含物变化研究[J]. 华北农学报, 2023, 38(S1): 253-260.
[14]	王江银, 徐婉宁, 苏洋, 张博. 干旱胁迫下紫花苜蓿和黄花苜蓿实生苗叶片形态及解剖结构变化[J]. 华北农学报, 2023, 38(S1): 228-236.
[15]	张曦, 刘祎, 钱玉源, 王广恩, 王寒菊, 焦秀芬, 解辉, 崔淑芳, 李俊兰. 陆地棉种质资源遗传多样性分析及初级核心种质构建[J]. 华北农学报, 2023, 38(S1): 26-33.

模态框（Modal）标题

选择文件类型/文献管理软件名称

选择包含的内容

GhLhcb2A1基因在陆地棉低温和干旱响应中的功能

Function Analysis of GhLhcb2A1 Gene in Response to Low Temperature and Drought in Upland Cotton

RichHTML

PDF

可视化

被引次数

摘要/Abstract

引用本文

使用本文

图/表 6

参考文献 32

相关文章 15

编辑推荐

Metrics

本文评价