响应NO的藜麦CHX基因家族在盐碱胁迫下的表达模式

doi:10.13560/j.cnki.biotech.bull.1985.2024-0500

摘要/Abstract

摘要：

目的 CHX（cation/H⁺ exchanger）基因家族是植物特有的一价阳离子转运蛋白，在植物的生长发育、盐碱胁迫响应中发挥重要作用。从藜麦全基因组范围内鉴定CHX基因家族成员，分析相关基因表达特点，为后续CHX基因功能深入研究提供支撑。方法利用生物信息学方法，从藜麦基因组中鉴定CHX基因家族成员，并对其理化特性、系统进化关系、染色体定位、基因复制、基因结构和启动子区顺式作用元件等进行分析。利用转录组数据结合RT-qPCR分析CHX家族基因在混合盐碱胁迫和外源NO处理下不同器官的表达模式。采用烟草瞬时转化法检测目标蛋白的亚细胞定位情况。结果在藜麦基因组中共鉴定到51个CHX基因，系统发育分析将CHX家族划分为5个亚族。片段复制是CHX基因家族进化的主要原因，该基因家族经历了强烈的纯化选择。在转录组分析基础上，对其中10个CHX基因在盐碱胁迫和NO处理下的RT-qPCR鉴定分析表明，除CqCHX-10和CqCHX-25不表达以外，其余8个基因均在茎和根系中响应盐碱胁迫，并受到外源NO的正向调控。进一步对CqCHX-17蛋白亚细胞定位发现，该基因定位于细胞核和叶绿体。结论从藜麦基因组中鉴定出51个CHX家族成员。不同成员在不同组织中表现出不同的表达模式。CHX基因家族成员对盐碱胁迫和外源NO正向响应，CqCHX-17可能参与K⁺转运，并参与叶绿体基质pH调节。

关键词: 藜麦, CHX, 生信分析, 表达分析, 盐碱胁迫

Abstract:

Objective CHX (cation/H⁺ exchanger) gene family is a unique monovalent cationic transporter and plays an important role in plant growth and development and response to salt-alkali stress. The CHX gene family members were identified from the whole genome of quinoa, and the expression characteristics of related genes were analyzed to provide support for further research on CHX gene function. Method Bioinformatics methods were used to identify the CHX gene family members from quinoa genome, and to analyze their physicochemical properties, phylogenetic relationships, chromosome localization, gene replication, gene structure and promoter cis-acting elements. Transcriptome data combined with RT-qPCR were used to analyze the expression patterns of CHX family genes in different organs under mixed saline-alkali stress and exogenous NO treatment. Transient transformation of tobacco was used to detect the subcellular localization of target protein. Result A total of 51 CHX genes were identified in quinoa genome, and the CHX family was divided into 5 subfamilies by phylogenetic analysis. Fragment replication was the primary reason for the evolution of the CHX gene family, which has undergone intense purification selection. Based on the transcriptomic analysis, RT-qPCR analysis of 10 CHX genes under salt-alkali stress and NO treatment showed that the other 8 genes were all in response to salt-alkali stress in stems and roots and were positively regulated by exogenous NO except CqCHX-10 and CqCHX-25 were not expressed. Further subcellular localization of CqCHX-17 protein showed that the gene was localized in the nucleus and chloroplast. Conclusion Fifty-one CHX family members are identified from quinoa genome. Different members show different expression patterns in different organizations. CHX gene family members respond positively to saline-alkali stress and exogenous NO. It is found that CqCHX-17 may be involved in K⁺ transport and pH regulation of chloroplast matrix.

Key words: quinoa, CHX, bioinformatics analysis, expression analysis, saline-alkali stress

贾子健, 王宝强, 陈立飞, 王义真, 魏小红, 赵颖. 响应NO的藜麦CHX基因家族在盐碱胁迫下的表达模式[J]. 生物技术通报, 2025, 41(2): 163-174.

JIA Zi-jian, WANG Bao-qiang, CHEN Li-fei, WANG Yi-zhen, WEI Xiao-hong, ZHAO Ying. Expression Patterns of CHX Gene Family in Quinoa in Response to NO under Saline-alkali Stress[J]. Biotechnology Bulletin, 2025, 41(2): 163-174.

图/表 8

参考文献 36

1	李霞, 刘传鑫, 徐彬, 等. 植物拒盐机制的研究进展 [J]. 中国农学通报, 2023, 39(27): 86-94.
	Li X, Liu CX, Xu B, et al. Plant salt-exclusion mechanism: a review [J]. Chin Agric Sci Bull, 2023, 39(27): 86-94.
2	Ismail A, Takeda S, Nick P. Life and death under salt stress: same players, different timing? [J]. J Exp Bot, 2014, 65(12): 2963-2979.
3	Sewelam N, Oshima Y, Mitsuda N, et al. A step towards understanding plant responses to multiple environmental stresses: a genome-wide study [J]. Plant Cell Environ, 2014, 37(9): 2024-2035.
4	Teakle NL, Tyerman SD. Mechanisms of Cl^-‍ transport contributing to salt tolerance [J]. Plant Cell Environ, 2010, 33(4): 566-589.
5	Chanroj S, Wang GY, Venema K, et al. Conserved and diversified gene families of monovalent cation/H⁺ antiporters from algae to flowering plants [J]. Front Plant Sci, 2012, 3: 25.
6	刘斌, 周子豪, 杨正利, 等. 茶树CHX基因家族全基因组鉴定及生物信息学分析[J]. 分子植物育种, 2023. .
	Liu B, Zhou ZH, Yang ZL, et al. Genome-wide identification of CHX gene family in tea plant (Camellia sinensis) and bioinformatics analysi[J]. Mol Plant Breed, 2023. .
7	Padmanaban S, Chanroj S, Kwak JM, et al. Participation of endomembrane cation/H⁺ exchanger AtCHX20 in osmoregulation of guard cells [J]. Plant Physiol, 2007, 144(1): 82-93.
8	Jia Q, Zheng C, Sun S, et al. The role of plant cation/proton antiporter gene family in salt tolerance [J]. Biol Plant, 2018, 62(4): 617-629.
9	Guan RX, Qu Y, Guo Y, et al. Salinity tolerance in soybean is modulated by natural variation in GmSALT3 [J]. Plant J, 2014, 80(6): 937-950.
10	Qi XP, Li MW, Xie M, et al. Identification of a novel salt tolerance gene in wild soybean by whole-genome sequencing [J]. Nat Commun, 2014, 5: 4340.
11	Ren SX, Lyle C, Jiang GL, et al. Soybean salt tolerance 1 (GmST1) reduces ROS production, enhances ABA sensitivity, and abiotic stress tolerance in Arabidopsis thaliana [J]. Front Plant Sci, 2016, 7: 445.
12	Jia BW, Sun MZ, DuanMu HZ, et al. GsCHX19.3, a member of cation/H⁺ exchanger superfamily from wild soybean contributes to high salinity and carbonate alkaline tolerance [J]. Sci Rep, 2017, 7(1): 9423.
13	Qu Y, Guan RX, Bose J, et al. GmSALT3 confers shoot Na⁺ and Cl^- exclusion in soybean via two distinct processes [J]. bioRxiv, 2020. DOI: 10.1101/2020.01.06.896456 .
14	Isayenkov SV, Maathuis FJM. Plant salinity stress: many unanswered questions remain [J]. Front Plant Sci, 2019, 10: 80.
15	Hall D, Evans AR, Newbury HJ, et al. Functional analysis of CHX21: a putative sodium transporter in Arabidopsis [J]. J Exp Bot, 2006, 57(5): 1201-1210.
16	Isayenkov SV, Dabravolski SA, Pan T, et al. Phylogenetic diversity and physiological roles of plant monovalent cation/H⁺ antiporters [J]. Front Plant Sci, 2020, 11: 573564.
17	Guo YQ, Zhu CR, Tian ZY. Overexpression of KvCHX enhances salt tolerance in Arabidopsis thaliana seedlings [J]. Curr Issues Mol Biol, 2023, 45(12): 9692-9708.
18	杨发荣, 黄杰, 魏玉明, 等. 藜麦生物学特性及应用 [J]. 草业科学, 2017, 34(3): 607-613.
	Yang FR, Huang J, Wei YM, et al. A review of biological characteristics, applications, and culture of Chenopodium quinoa [J]. Pratacultural Sci, 2017, 34(3): 607-613.
19	Vilcacundo R, Hernández-Ledesma B. Nutritional and biological value of quinoa (Chenopodium quinoa Willd.) [J]. Curr Opin Food Sci, 2017, 14: 1-6.
20	王艺臻, 丁国栋, 崔欣然, 等. 盐碱复合胁迫对油沙豆生长和光合特性的影响 [J]. 干旱区资源与环境, 2022, 36(5): 146-152.
	Wang YZ, Ding GD, Cui XR, et al. Effects of saline-alkali stress on the growth and photosynthetic characteristics of Cyperus esculentus and the responses of protective enzymes [J]. J Arid Land Resour Environ, 2022, 36(5): 146-152.
21	Jarvis DE, Ho YS, Lightfoot DJ, et al. The genome of Chenopodium quinoa [J]. Nature, 2017, 542(7641): 307-312.
22	彭悦, 丁洪霞, 杨博, 等. 藜麦CqNHX基因家族表达与耐盐相关性分析[J/OL]. 分子植物育种, 2023. .
	Peng Y, Ding HX, Yang B, et al. Identification and expression analysis of CqNHX gene family in quinoa under salt stress [J/OL]. Mol Plant Breed, 2023. .
23	褚晶. 藜麦GST基因家族的鉴定与表达分析及发根农杆菌介导的GST基因功能研究 [D]. 烟台: 烟台大学, 2022.
	Chu J. Identification and expression analysis of GST gene family in quinoa and study on GST gene function mediated by Agrobacterium rhizogenes [D]. Yantai: Yantai University, 2022.
24	Zhu XL, Wang BQ, Wei XH. Identification and expression analysis of the CqSnRK2 gene family and a functional study of the CqSnRK2.12 gene in quinoa (Chenopodium quinoa Willd.) [J]. BMC Genomics, 2022, 23(1): 397.
25	Sze H, Padmanaban S, Cellier F, et al. Expression patterns of a novel AtCHX gene family highlight potential roles in osmotic adjustment and K⁺ homeostasis in pollen development [J]. Plant Physiol, 2004, 136(1): 2532-2547.
26	贾博为, 金军, 庄齐, 等. 玉米CHX基因家族全基因组鉴定与表达分析 [J]. 黑龙江八一农垦大学学报, 2022, 34(4): 23-30, 64.
	Jia BW, Jin J, Zhuang Q, et al. Genome-wide identification and expression analysis of maize CHX family [J]. J Heilongjiang Bayi Agric Univ, 2022, 34(4): 23-30, 64.
27	才晓溪, 沈阳, 周伍红, 等. 大豆CHX基因家族全基因组鉴定与生物信息学分析 [J]. 基因组学与应用生物学, 2018, 37(12): 5360-5369.
	Cai XX, Shen Y, Zhou WH, et al. Genome-wide identification and bioinformatics analysis of soybean CHX gene family [J]. China Ind Econ, 2018, 37(12): 5360-5369.
28	Abdullah, Faraji S, Mehmood F, et al. The GASA gene family in cacao (Theobroma cacao, Malvaceae): genome wide identification and expression analysis [J]. Agronomy, 2021, 11(7): 1425.
29	Musavizadeh Z, Najafi-Zarrini H, Kazemitabar SK, et al. Genome-wide analysis of Potassium channel genes in rice: expression of the OsAKT and OsKAT genes under salt stress [J]. Genes, 2021, 12(5): 784.
30	Heidari P, Faraji S, Ahmadizadeh M, et al. New insights into structure and function of TIFY genes in Zea mays and Solanum lycopersicum: a genome-wide comprehensive analysis [J]. Front Genet, 2021, 12: 657970.
31	Faraji S, Heidari P, Amouei H, et al. Investigation and computational analysis of the sulfotransferase (SOT) gene family in potato (Solanum tuberosum): insights into sulfur adjustment for proper development and stimuli responses [J]. Plants, 2021, 10(12): 2597.
32	吴彤, 刘云苗, 金军, 等. 蒺藜苜蓿cation/H⁺ exchanger基因家族鉴定及表达特征分析 [J]. 草业学报, 2022, 31(1): 181-194.
	Wu T, Liu YM, Jin J, et al. Identification and expression characteristics of a cation/H⁺ exchanger gene family in Medicago truncatula [J]. Acta Prataculturae Sin, 2022, 31(1): 181-194.
33	Pardo JM, Cubero B, Leidi EO, et al. Alkali cation exchangers: roles in cellular homeostasis and stress tolerance [J]. J Exp Bot, 2006, 57(5): 1181-1199.
34	Haro R, Fraile-Escanciano A, González-Melendi P, et al. The potassium transporters HAK2 and HAK3 localize to endomembranes in Physcomitrella patens. HAK2 is required in some stress conditions [J]. Plant Cell Physiol, 2013, 54(9): 1441-1454.
35	Sze H, Chanroj S. Plant endomembrane dynamics: studies of K⁺/H⁺ antiporters provide insights on the effects of pH and ion homeostasis [J]. Plant Physiol, 2018, 177(3): 875-895.
36	Song CP, Guo Y, Qiu QS, et al. A probable Na⁺(K⁺)/H⁺ exchanger on the chloroplast envelope functions in pH homeostasis and chloroplast development in Arabidopsis thaliana [J]. Proc Natl Acad Sci U S A, 2004, 101(27): 10211-10216.

基因 Gene	正向引物 Forward primer （5′-3′）	反向引物 Reverse primer （5′-3′）
CqCHX-6	TGACTGGTGTGATTGCTCCC	GTTGCTGGCTTCCAGGAGAT
CqCHX-10	CAGCGCTGGTGATGAGAGAT	TCTTCCTGAGTCGGCAAACC
CqCHX-16	TCCATGTGCATTGCTGTTGC	AGAAGAATCCAGGCTGCCAC
CqCHX-17	GTCCACTTCAGAGCAGACCC	CATCAGGTGCCTTGGAGTGT
CqCHX-18	GTGAGCCAGTGGATGAGCTT	CAAATGCACCGAAAAGGGCA
CqCHX-22	GAACGCTTTGCCAACACCAT	TGATCGCCTTCCTCCCTGTA
CqCHX-24	CAACCTCCACGGGAGTCTTC	GAACACCTCGCTACGACCAA
CqCHX-25	GTTAGCCAAGGCCCCTTCAT	CATGGCCATACGGCCTACAT
CqCHX-26	GGCATCACATTGCCGTTTGT	ATAGAGAGGGCAACCCCCAT
CqCHX-48	GGGTTTACTCGTGCTGGTGA	GGACTAACGCATCCCTAGCC
CqTUB-9	GAGATGTTCCGTCGTGTGAGTGAG	ATCGGCAGTTGCATCCTGGTATTG

基因 Gene	正向引物 Forward primer （5′-3′）	反向引物 Reverse primer （5′-3′）
CqCHX-6	TGACTGGTGTGATTGCTCCC	GTTGCTGGCTTCCAGGAGAT
CqCHX-10	CAGCGCTGGTGATGAGAGAT	TCTTCCTGAGTCGGCAAACC
CqCHX-16	TCCATGTGCATTGCTGTTGC	AGAAGAATCCAGGCTGCCAC
CqCHX-17	GTCCACTTCAGAGCAGACCC	CATCAGGTGCCTTGGAGTGT
CqCHX-18	GTGAGCCAGTGGATGAGCTT	CAAATGCACCGAAAAGGGCA
CqCHX-22	GAACGCTTTGCCAACACCAT	TGATCGCCTTCCTCCCTGTA
CqCHX-24	CAACCTCCACGGGAGTCTTC	GAACACCTCGCTACGACCAA
CqCHX-25	GTTAGCCAAGGCCCCTTCAT	CATGGCCATACGGCCTACAT
CqCHX-26	GGCATCACATTGCCGTTTGT	ATAGAGAGGGCAACCCCCAT
CqCHX-48	GGGTTTACTCGTGCTGGTGA	GGACTAACGCATCCCTAGCC
CqTUB-9	GAGATGTTCCGTCGTGTGAGTGAG	ATCGGCAGTTGCATCCTGGTATTG

重复CHX基因 Duplicated CHX gene	非同义替换率 Ka	同义替换率 Ks	比值 Ka/Ks	重复CHX基因 Duplicated CHX gene	非同义替换率 Ka	同义替换率 Ks	比值 Ka/Ks
CqCHX-1-CqCHX-11	0.015	0.106	0.140	CqCHX-17-CqCHX-24	0.092	0.471	0.196
CqCHX-2-CqCHX-33	0.143	0.781	0.183	CqCHX-17-CqCHX-26	0.111	0.615	0.181
CqCHX-4-CqCHX-13	0.057	0.182	0.316	CqCHX-18-CqCHX-26	0.032	0.166	0.191
CqCHX-4-CqCHX-5	0.111	0.296	0.376	CqCHX-18-CqCHX-25	0.104	0.610	0.170
CqCHX-6-CqCHX-22	0.004	0.060	0.066	CqCHX-18-CqCHX-41	0.119	0.527	0.225
CqCHX-7-CqCHX-30	0.021	0.171	0.126	CqCHX-19-CqCHX-34	0.015	0.193	0.080
CqCHX-8-CqCHX-29	0.032	0.159	0.203	CqCHX-21-CqCHX-50	0.085	0.151	0.566
CqCHX-9-CqCHX-28	0.018	0.163	0.112	CqCHX-24-CqCHX-25	0.076	0.515	0.148
CqCHX-14-CqCHX-45	0.012	0.169	0.073	CqCHX-24-CqCHX-41	0.096	0.464	0.207
CqCHX-15-CqCHX-44	0.012	0.116	0.106	CqCHX-24-CqCHX-26	0.116	0.583	0.200
CqCHX-16-CqCHX-24	0.028	0.122	0.230	CqCHX-25-CqCHX-41	0.047	0.323	0.146
CqCHX-16-CqCHX-25	0.080	0.520	0.154	CqCHX-25-CqCHX-26	0.103	0.600	0.172
CqCHX-16-CqCHX-41	0.101	0.483	0.210	CqCHX-26-CqCHX-41	0.115	0.599	0.192
CqCHX-17-CqCHX-25	0.030	0.198	0.153	CqCHX-39-CqCHX-40	0.022	0.144	0.151
CqCHX-17-CqCHX-41	0.064	0.333	0.191

重复CHX基因 Duplicated CHX gene	非同义替换率 Ka	同义替换率 Ks	比值 Ka/Ks	重复CHX基因 Duplicated CHX gene	非同义替换率 Ka	同义替换率 Ks	比值 Ka/Ks
CqCHX-1-CqCHX-11	0.015	0.106	0.140	CqCHX-17-CqCHX-24	0.092	0.471	0.196
CqCHX-2-CqCHX-33	0.143	0.781	0.183	CqCHX-17-CqCHX-26	0.111	0.615	0.181
CqCHX-4-CqCHX-13	0.057	0.182	0.316	CqCHX-18-CqCHX-26	0.032	0.166	0.191
CqCHX-4-CqCHX-5	0.111	0.296	0.376	CqCHX-18-CqCHX-25	0.104	0.610	0.170
CqCHX-6-CqCHX-22	0.004	0.060	0.066	CqCHX-18-CqCHX-41	0.119	0.527	0.225
CqCHX-7-CqCHX-30	0.021	0.171	0.126	CqCHX-19-CqCHX-34	0.015	0.193	0.080
CqCHX-8-CqCHX-29	0.032	0.159	0.203	CqCHX-21-CqCHX-50	0.085	0.151	0.566
CqCHX-9-CqCHX-28	0.018	0.163	0.112	CqCHX-24-CqCHX-25	0.076	0.515	0.148
CqCHX-14-CqCHX-45	0.012	0.169	0.073	CqCHX-24-CqCHX-41	0.096	0.464	0.207
CqCHX-15-CqCHX-44	0.012	0.116	0.106	CqCHX-24-CqCHX-26	0.116	0.583	0.200
CqCHX-16-CqCHX-24	0.028	0.122	0.230	CqCHX-25-CqCHX-41	0.047	0.323	0.146
CqCHX-16-CqCHX-25	0.080	0.520	0.154	CqCHX-25-CqCHX-26	0.103	0.600	0.172
CqCHX-16-CqCHX-41	0.101	0.483	0.210	CqCHX-26-CqCHX-41	0.115	0.599	0.192
CqCHX-17-CqCHX-25	0.030	0.198	0.153	CqCHX-39-CqCHX-40	0.022	0.144	0.151
CqCHX-17-CqCHX-41	0.064	0.333	0.191

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