Abstract:Plant cyclic nucleotide-gated ion channel (CNGC) plays an important role in plant growth, development and stress response. In order to investigate the function of CNGC gene family in pepper (Capsicum annuum), the CNGC family members of pepper were identified, and the physical and chemical properties, subcellular localization, phylogenetic development, structural characteristics, promoter cis-acting elements and gene expression patterns were analyzed. The results showed that a total of 14 CaCNGC gene family members were identified in pepper, which were not uniformly distributed on each chromosome. The isoelectric point of CaCNGC was 8.64~9.61, the molecular weight was 72.87~91.49 kD. The instability index was greater than 40, indicating that 14 members of CaCNGC were unstable proteins. All CaCNGC proteins were located in the plasma membrane, had transmembrane structures, and were hydrophilic proteins. The analysis of cis-element in promoter showed that, most of the CaCNGC contained cis-elements related to growth and development, hormone regulation, and abiotic stresses response. Transcriptome data analysis showed that most members of CaCNGC had low or no expression in leaves, flowers and fruits of pepper. Most members of CaCNGC were expressed under low temperature, high temperature, abscisic acid, gibberellin, hydrogen peroxide, indoleacetic acid, jasmonic acid and salt stress. qRT-PCR analysis showed that the expression of CaCNGC2, CaCNGC7 and CaCNGC10, was up-regulated under low and high temperature stress, which might be involved in the response of pepper to temperature stress. These results provide a theoretical reference for further investigating the function of CaCNGC gene family, and new insights into the molecular mechanisms of pepper tolerance to temperature stress.
王田田, 常雪瑞, 王静. 辣椒CNGC基因家族鉴定及表达分析[J]. 农业生物技术学报, 2025, 33(2): 271-284.
WANG Tian-Tian, CHANG Xue-Rui, WANG Jing. Identification and Expression Analysis of CNGC Gene Family in Pepper (Capsicum annuum). 农业生物技术学报, 2025, 33(2): 271-284.
[1] 邓惠如, 张素勤, 耿广东. 2023. 辣椒逆境响应基因研究现状及前景[J/OL]. 分子植物育种, 1-12. https://link.cnki.net/urlid/46.1068.S.20231108.1149.002. (Deng H R, Zhang S Q, Geng G D. 2023. Research status and prospect of stress response genes in pepper[J/OL]. Molecular Plant Breeding, 1-12. https://link.cnki.net/urlid/46.1068.S.20231108.1149.002. [2] 耿小惠. 2022. 钙离子通道促进剂及抑制剂对香蕉抗寒性的影响及相关基因MaCNGC家族成员鉴定与表达分析[D]. 硕士学位论文, 福建农林大学, 导师: 赖钟雄, 林文忠, pp. 3-34. (Geng X H. 2022. Effect of calcium channel promoters and inhibitors on cold resistance in banana and identification and expression analysis of MaCNGC family members[D]. Thesis for M.S., Fujian Agriculture and Forestry University, Supervisor: Lai Z X, Lin W Z, pp. 3-34.) [3] 郝宇瑞, 赖长巍, 刘恋, 等. 2023. 马铃薯CNGC家族全基因组鉴定和组织特异性分析[J]. 分子植物育种, 03: 1-14. (Hao Y R, Lai C W, Liu L, et al. 2023. Identification and analysis of CNGC family genes in potato (Solanum tuberosum)[J]. Molecular Plant Breeding, 03: 1-14.) [4] 刘文豪, 田琴, 余渝, 等. 2021. 陆地棉CNGC全基因组鉴定及表达分析[J]. 江苏农业科学, 49(24): 49-56. (Liu W H, Tian Q, Yu Y, et al. 2021. Genome-wide identification and expression analysis of Gossypium hirsutum Linn. CNGC[J]. Jiangsu Agricultural Sciences, 49(24): 49-56.) [5] 刘震, 陈丽敏, 李志涛,等. 2024. 马铃薯ARM基因家族的全基因组鉴定及表达分析[J]. 作物学报, 50 (06): 1451-1466. (Liu Z, Chen L M, Li Z H, et al. 2024. Genome-wide identification and expression analysis of ARM gene family inpotato (Solanum tuberosum L.)[J]. Acta Agronomica Sinica, 2024, 50(6): 1451-1466.) [6] 马艳青, 戴雄泽, 李雪峰, 等. 2015. 辣椒骨干亲本6421及其衍生系的创制与利用[J]. 中国蔬菜, 06: 11-16. (Ma Y Q, Dai X Z, Li X F, et al. 2015. Creation and utilization of pepper backbone parent 6421 and its derivatives[J]. Chinese Vegetables, 06: 11-16.) [7] 马玉虎, 魏敏, 段盼盼, 等. 2023. 辣椒PP2C家族基因鉴定与响应低温胁迫的表达分析[J]. 干旱地区农业研究, 41(06): 39-53. (Ma Y H, Wei M, Duan P P, et al. 2023. Identification and expression analysis of PP2C family genes in pepper in response to low temperature stress[J]. Agricultural Research in Drylands, 41(06): 39-53.) [8] 田韦韦, 严志祥, 王成,等. 2023. 基于全长转录组测序的多花黄精WRKY转录因子家族分析[J]. 中国中药杂志, 48 (04): 939-950. (Tin W W, Yan Z X, Wang C, et al. 2023. Analysis of WRKY transcription factor family based on full-lengthtranscriptome sequencing in Polygonatum cyrtonema[J]. China Journal of Chinese Materia, 48 (04): 939-950.) [9] 王静, 谭放军, 梁成亮, 等. 2020. 辣椒热激蛋白HSP90家族基因鉴定及分析[J]. 园艺学报, 47(04): 665-674. (Wang J, Tan F J, Liang C L, et al. 2020. Genome-wide identification and analysis of HSP90 gene family in pepper[J]. Journal of Horticulture, 47(04): 665-674.) [10] 王立浩, 张宝玺, 张正海, 等. 2021. “十三五”我国辣椒育种研究进展、产业现状及展望[J]. 中国蔬菜, (02): 21-29. (Wang L H, Zhang B X, Zhang Z H, et al. 2021. Status in breeding and production of Capsicum spp. in China during 'the thirteenth five-year plan' period and future prospect[J]. China Vegetables, (02): 21-29.) [11] 王欣悦, 刘培源, 余冰清, 等. 2021. 植物环核苷酸门控离子通道的研究进展[J]. 湖北农业科学, 60(08): 5-11. (Wang X Y, Liu P Y, Yu B Q, et al. 2021. Research advances in plant cyclic nucleotide-gated channels[J]. Hubei Agricultural Sciences, 60(08): 5-11.) [12] 王雪莹, 王瑞琪, 张洋, 等. 2022. 毛果杨CNGC家族全基因组鉴定及胁迫响应分析[J]. 植物研究, 42(04): 613-625. (Wang X Y, Wang R Q, Zhang Y, et al. 2022. Whole genome identification and stress response analysis of the CNGC family of Populus pilocarpa[J]. Plant Research, 42(04): 613-625.) [13] 邹学校, 朱凡. 2022. 辣椒的起源、进化与栽培历史[J]. 园艺学报, 49(06): 1371-1381. (Zou X X, Zhu F. 2022. Origin, evolution and cultivation history of the pepper[J]. Acta Horticulturae Sinica, 49(06): 1371-1381.) [14] Bailey T L, Johnson J, Grant C E, et al. 2015. The MEME Suite[J]. Nucleic Acids Research, 43(W1): W39-W49. [15] Chao J T, Kong Y Z, Wang Q, et al. 2015. MapGene2Chrom, a tool to draw gene physical map based on Perl and SVG languages[J]. Yi Chuan, 37(1): 91-97. [16] Chen C J, Wu Y, Li J W, et al. 2023. TBtools-II: A "one for all, all for one" bioinformatics platform for biological big-data mining[J]. Molecular Plant, 16(11): 1733-1742. [17] Chen J Q, Yin H, Gu J P, et al. 2015. Genomic characterization, phylogenetic comparison and differential expression of the cyclic nucleotide-gated channels gene family in pear (Pyrus bretchneideri Rehd.)[J]. Genomics, 105(1): 39-52. [18] Chen L, Wang W W, He H L, et al. 2023. Genome-wide identification, characterization and experimental expression analysis of CNGC gene family in Gossypium[J]. International Journal of Molecular Sciences, 24(5): 4617. [19] Cui Y M, Lu S, Li Z, et al. 2020. Cyclic nucleotide-gated ion channels 14 and 16 promote tolerance to heat and chilling in rice[J]. Plant Physiology, 183(4): 1794-1808. [20] Cui Y X, Wang J X, Bai Y X, et al. 2023. Identification of CNGCs in glycine max and screening of related resistance genes after Fusarium solani infection[J]. Biology-Basel, 12(3): 439. [21] Dodd A N, Gardner M J, Hotta C T, et al. 2007. The Arabidopsis circadian clock incorporates a cADPR-based feedback loop[J]. Science. 318(5857): 1789-1792. [22] Finn R D, Clements J, Eddy S R. 2011. HMMER web server: Interactive sequence similarity searching[J]. Nucleic Acids Research, 39: W29-W37. [23] Frietsch S, Wang Y F, Sladek C, et al. 2007. A cyclic nucleotide-gated channel is essential for polarized tip growth of pollen[J]. Proceedings of the National Academy of Sciences of the USA, 104(36): 14531-14536. [24] Gao F, Han X W, Wu J H, et al. 2012. A heat-activated calcium-permeable channel Arabidopsis cyclic nucleotide-gated ion channel 6 is involved in heat shock responses[J]. Plant Journal, 70(6): 1056-1069. [25] Guo J, Islam M A, Lin H C, et al. 2018. Genome-wide identification of cyclic nucleotide-gated ion channel gene family in wheat and functional analyses of TaCNGC14 and TaCNGC16[J]. Frontiers in Plant Science, 9: 18. [26] Hao L D, Qiao X L. 2018. Genome-wide identification and analysis of the CNGC gene family in maize[J]. PeerJ, 6: e5816. [27] Harada A, Sakai T, Okada K. 2003. Phot1 and phot2 mediate blue light-induced transient increases in cytosolic Ca2+ differently in Arabidopsis leaves[J]. Proceedings of the National Academy of Sciences of the USA, 100(14): 8583-8588. [28] Hu B, Jin J, Guo A Y, et al. 2015. GSDS 2.0: An upgraded gene feature visualization server[J]. Bioinformatics, 31(8): 1296-1297. [29] Jarratt-Barnham E, Wang L, Ning Y, et al. 2021. The complex story of plant cyclic nucleotide-gated channels[J]. International Journal of Molecular Sciences, 22(2): 874. [30] Jiang Z, Du L H, Shen L, et al. 2023. Genome-wide exploration and expression analysis of the CNGC gene family in eggplant (Solanum melongena L.) under cold stress, with functional characterization of SmCNGC1a[J]. International Journal of Molecular Sciences, 24(17): 13049. [31] Kakar K U, Nawaz Z, Kakar K, et al. 2017. Comprehensive genomic analysis of the CNGC gene family in Brassica oleracea: Novel insights into synteny, structures, and transcript profiles[J]. BMC Genomics, 18(1): 869. [32] Kosuta S, Hazledine S, Sun J, et al. 2008. Differential and chaotic calcium signatures in the symbiosis signaling pathway of legumes[J]. Proceedings of the National Academy of Sciences of the USA, 105(28): 9823-9828. [33] Kumar S, Stecher G, Li M, et al. 2018. MEGA X: Molecular evolutionary genetics analysis across computing platforms[J]. Molecular Biology and Evolution. 35(6): 1547-1549. [34] Lescot M, Déhais P, Thijs G, et al. 2002. PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences[J]. Nucleic Acids Research, 30(1): 325-327. [35] Li Q Q, Yang S Q, Ren J, et al. 2019. Genome-wide identification and functional analysis of the cyclic nucleotide-gated channel gene family in Chinese cabbage[J]. 3 Biotech, 9(3): 114. [36] Liu F, Yu H, Deng Y, et al. 2017. PepperHub, an informatics hub for the chili pepper research community[J]. Molecular Plant, 10(8): 1129-1132. [37] Liu Z, Song J, Miao W, et al. 2021. Comprehensive proteome and lysine acetylome analysis reveals the widespread involvement of acetylation in cold resistance of pepper (Capsicum annuum L.)[J]. Frontiers in Plant Science, 12: 730489. [38] M?ser P, Thomine S, Schroeder J I, et al. 2001. Phylogenetic relationships within cation transporter families of Arabidopsis[J]. Plant Physiology, 126(4): 1646-1667. [39] Mistry J, Chuguransky S, Williams L, et al. 2021. Pfam: The protein families database in 2021[J]. Nucleic Acids Research, 49(D1): D412-D419. [40] M?ller S, Croning M D, Apweiler R. 2002. Evaluation of methods for the prediction of membrane spanning regions[J]. Bioinformatics, 18(1): 218. [41] Nawaz Z, Kakar K U, Saand M A, et al. 2014. Cyclic nucleotide-gated ion channel gene family in rice, identification, characterization and experimental analysis of expression response to plant hormones, biotic and abiotic stresses[J]. BMC Genomics, 15(1): 853. [42] Nawaz Z, Kakar K U, Ullah R, et al. 2019. Genome-wide identification, evolution and expression analysis of cyclic nucleotide-gated channels in tobacco (Nicotiana tabacum L.)[J]. Genomics, 111(2): 142-158. [43] Nielsen H. 2017. Predicting Secretory Proteins with SignalP[J]. Methods in Molecular Biology, 1611: 59-73. [44] Qi Z, Verma R, Gehring C, et al. 2010. Ca2+ signaling by plant Arabidopsis thaliana Pep peptides depends on AtPepR1, a receptor with guanylyl cyclase activity, and cGMP-activated Ca2+ channels[J]. Proceedings of the National Academy of Sciences of the USA, 107(49): 21193-21198. [45] Saand M A, Xu Y P, Li W, et al. 2015. Cyclic nucleotide gated channel gene family in tomato: Genome-wide identification and functional analyses in disease resistance[J]. Frontiers in Plant Science, 6: 303. [46] Saand M A, Xu Y P, Munyampundu J P, et al. 2015. Phylogeny and evolution of plant cyclic nucleotide-gated ion channel (CNGC) gene family and functional analyses of tomato CNGCs[J]. DNA Research, 22(6): 471-483. [47] Schuurink R C, Shartzer S F, Fath A, et al. 1998. Characterization of a calmodulin-binding transporter from the plasma membrane of?barley?aleurone[J]. Proceedings of the National Academy of Sciences of the USA, 95(4): 1944-1949. [48] Srivastava A, Singh K, Khar A, et al. 2022. Morphological, biochemical and molecular insights on responses to heat stress in chilli[J]. Indian Journal of Horticulture, 79(1): 15-22. [49] Thompson J D, Gibson T J, Higgins D G. 2002. Multiple sequence alignment using ClustalW and ClustalX[J]. Current Protocols in Bioinformatics, 2: Unit 2.3. [50] Tracy F E, Gilliham M, Dodd A N, et al. 2008. NaCl-induced changes in cytosolic free Ca2+ in Arabidopsis thaliana are heterogeneous and modified by external ionic composition [J]. Plant Cell and Environment, 31(8): 1063-1073. [51] Wang J C, Liu X, Zhang A, et al. 2019. A cyclic nucleotide-gated channel mediates cytoplasmic calcium elevation and disease resistance in rice[J]. Cell Research, 29(10): 820-831. [52] Wang L X, Li M, Liu Z G, et al. 2020. Genome-wide identification of CNGC genes in Chinese jujube (Ziziphus jujuba Mill.) and ZjCNGC2 mediated signalling cascades in response to cold stress[J]. BMC Genomics, 21(1): 191. [53] Waterhouse A, Bertoni M, Bienert S, et al. 2018. SWISS-MODEL: Homology modelling of protein structures and complexes[J]. Nucleic Acids Research, 46(W1): W296-W303. [54] White P J. 2000. Calcium channels in higher plants[J]. Biochimica et Biophysica Acta-Biomembranes, 1465(1-2): 171-189. [55] Wilkins M R, Gasteiger E, Bairoch A, et al. 1999. Protein identification and analysis tools in the ExPASy server[J]. Methods in Molecular Biology, 112: 531-552. [56] Zhang N N, Lin H Z, Zeng Q Y, et al. 2023. Genome-wide identification and expression analysis of the cyclic nucleotide-gated ion channel (CNGC) gene family in Saccharum spontaneum[J]. BMC Genomics, 24(1): 281. [57] Zhang Y J, Li Y B, Yang J, et al. 2023. Genome-wide analysis and expression of cyclic nucleotide-gated ion channel (CNGC) family genes under cold stress in Mango (Mangifera indica)[J]. Plants-Basel, 12(3): 592. [58] Zia K, Rao M J, Sadaqat M, et al. 2022. Pangenome-wide analysis of cyclic nucleotide-gated channel (CNGC) gene family in Citrus spp. revealed their intraspecies diversity and potential roles in abiotic stress tolerance[J]. Frontiers in Genetics, 13: 1034921.