1 College of Life Sciences/Institute of Agro-bioengineering/Key Laboratory of Plant Resources Conservation and Germplasm Innovationin in Mountainous Region (Ministry of Education), Guizhou University, Guiyang 550025, China; 2 College of Tea Sciences, Guizhou University, Guiyang 550025, China; 3 Guizhou Academy of Agricultural Sciences, Guiyang 550006, China
Abstract:Tea plants (Camellia sinensis), as one of the important woody economic plants, are vulnerable to low temperature and freezing injury, which seriously affecting the yield and quality of tea. In order to further clarify the molecular mechanism on cold response of tea plants, in this study, based on the cold treatment transcriptome library of 2 Guizhou cultivars, Qiancha 1 (QC1) and Qianmei 601 (QM601), a cold-upregulated differentially expressed mitogen-activated protein kinase (MAPK) gene (transcriptome library number: CSS0027011) was isolated and cloned, and named as CsMAPK15 (GenBank No. ON39909). The open reading frame of CsMAPK15 contains 1 701 nucleotides encoding a protein of 566 amino acids. Motif analysis revealed that CsMAPK15 had 3 conserved motifs. Phylogenetic analysis showed that CsMAPK15 shared high sequence similarity with homologs from other species. It shared 79.17% identity with AtMAPK16 (AT5G19010) from Arabidopsis and 80.51 % similarity with OsMAPK15 (LOC_Os07g47490) from rice (Oryza sativa). The tissue pattern analysis revealed that expression level of CsMAPK15 was highest in roots. Promoter analysis showed that the CsMAPK15 promoter had several cis-acting elements, including stress responsive element (STRE) and salicylic acid (SA) responsive element (TCA element). qPCR confirmed that CsMAPK15 could response to cold signal and SA. Furthermore, the transgenic rice plants which overexpressed CsMAPK15 were successfully obtained by callus transformation. The preliminary results showed that there was no significant change in plant growth and development between transgenic plants and wild-type plants. This study provides research materials and theoretical basis for further clarifying the role of CsMAPK15 in plant cold resistance regulation.
[1] 曹红利, 郝心愿, 岳川, 等. 2013. 茶树甜菜碱醛脱氢酶基因(CsBADH1)的全长cDNA克隆与表达分析[J]. 茶叶科学, 33(02): 99-108. (Cao H L, Hao X Y, Yue C, et al.2013. Cloning and expression analysis of betaine aldehyde dehydrogenase gene (CsBADH1) from tea plant[J]. Journal of Tea Science, 33(02): 99-108.) [2] 崔永祯, 赵红, 黄格, 等. 2020. 农杆菌介导的冷诱导基因水稻遗传转化体系的建立[J]. 江西农业学报, 32(07): 6-11. (Cui Y Z, Zhao H, Huang G, et al.2020. Establishment of genetic transformation system of cold-induced gene mediated by Agrobacterium in rice[J]. Acta Agricultural Jiangxi, 32(07): 6-11.) [3] 梁少奎, 阳国梁, 胡慧, 等. 2022. 虾青素通过激活MAPK信号通路诱导睾丸生殖细胞瘤细胞凋亡和自噬[J]. 生命科学研究, 26(01): 39-46. (Liang S K, Yang G L, Hu H, et al.2022. Astaxanthin induces apoptosis and autophagy of testicular germ cell tumor cells via activating MAPK signaling pathway[J]. Life Science Research, 26(01): 39-46.) [4] 刘亚菲, 张帆, 梁卫红. 2021. 水稻MAPK级联的功能和作用机制[J]. 中国生物化学与分子生物学报, 37(12): 1569-1576. (Liu Y F, Zhang F, Liang W H.2021. Function and mechanism of MAPK cascades in rice[J]. Chinese Journal of Biochemistry and Molecular Biology, 37(12): 1569-1576.) [5] 吴红姣. 2020. MAPK信号级联及下游基质金属蛋白酶在拟南芥叶片衰老中的作用及调控机制研究[D]. 博士学位论文, 浙江大学, 导师: 张舒群, pp: 25-44. (Wu H J.2020. Regulation of Arabidopsis matrix metalloproteinases by MAP kinases and their function in leaf senescence[D]. Thesis for Ph. D., Zhejiang University, Supervisor: Zhang S Q, pp. 25-44.) [6] 杨春, 郭燕, 周顺珍, 等. 2015. 3个茶树品种(品系)鲜叶香气特征及萜烯指数分析[J]. 信阳师范学院学报(自然科学版), 28(04): 501-506. (Yang C, Guo Y, Zhou S Z, et al.2015. Analysis of volatile aroma components and terpene index in 3 tea varieties or strains[J]. Journal of Xinyang Normal University (Natural Science Edition), 28(04): 501-506.) [7] 叶新如, 王彬, 陈敏氡, 等. 2020. 冬瓜BhMAPK15基因的克隆及其非生物胁迫下的表达分析[J]. 中国细胞生物学学报, 42(09): 1526-1537. (Ye X R, Wang B, Chen N D, et al.2020. Cloning and abiotic stress expression analysis of BhMAPK15 gene in Benincasa hispida (Thunb.) Cogn[J]. Chinese Journal of Cell Biology, 42(09): 1526-1537.) [8] Ahlfors R, Macioszer V, Rudd J, et al.2004. Stress hormone-independent activation and nuclear translocation of mitogen-activated protein kinases in Arabidopsis thaliana during ozone exposure[J]. The Plant Journal, 40(4): 512-522. [9] Chatterjee A, Paul A, Unnati G M, et al.2020. MAPK cascade gene family in Camellia sinensis: In-silico identification, expression profiles and regulatory network analysis[J]. BMC Genomics, 21(01): 613-629. [10] Ding C, Lei L, Yao L, et al.2019. The involvements of calcium-dependent protein kinases and catechins in tea plant[Camellia sinensis (L.) O. Kuntze] cold responses[J]. Plant Physiology and Biochemistry, 143: 190-202. [11] Ding Y, Yang S.2022. Surviving and thriving: How plants perceive and respond to temperature stress[J]. Developmental Cell, 57(08): 947-958. [12] Ding Z T, Li C, Shi H, et al.2015. Pattern of CsICE1 expression under cold or drought treatment and functional verification through analysis of transgenic Arabidopsis[J]. Genetics and Molecular Research, 14(03): 11259-11270. [13] Hamel L, Nicole M, Sritubtim S, et al.2006. Ancient signals: Comparative genomics of plant MAPK and MAPKK gene families[J]. Trends in Plant Science, 11(04): 192-198. [14] Hao X, Tang H, Wang B.2018. Integrative transcriptional and metabolic analyses provide insights into cold spell response mechanisms in young shoots of the tea plant[J]. Tree Physiology, 38(11): 1-17. [15] Hong Y, Liu Q, Cao Y, et al.2019. The OsMAPK15 negatively regulates Magnaporthe oryza and xoo disease resistance via SA and JA signaling pathway in rice[C]. Frontiers in Plant Science, 10: 752. [16] Hu Y, Zhang M, Lu M, et al.2022. Salicylic acid carboxyl glucosyltransferase UGT87E7 regulates disease resistance in Camellia sinensis[J]. Plant Physiology, 188(3): 1507-1520. [17] Kumar K, Raina S, Sultan S, et al.2020. Arabidopsis MAPK signaling pathways and their cross talks in abiotic stress response[J]. Journal of Plant Biochemistry and Biotechnology, 29(4): 700-714. [18] Laloi C, Apel K, Danon A.2004. Reactive oxygen signalling: The latest news[J]. Current Opinion in Plant Biology, 7(3): 323-328. [19] Li X, Feng Z, Yang H, et al.2010. A novel cold-regulated gene from Camellia sinensis, CsCOR1, enhances salt-and dehydration-tolerance in tobacco[J]. Biochemical and Biophysical Research Communications, 394(02): 354-359. [20] Li Y, Zhang Q, Ou L, et al.2020. Response to the cold stress signaling of the tea plant (Camellia sinensis) elicited by chitosan oligosaccharide[J]. Agronomy Journal, 10(06): 915-926. [21] Mittler R, Kim Y, Song L, et al.2006. Gain-and loss-of-function mutations in Zat10 enhance the tolerance of plants to abiotic stress[J]. FEBS Letters, 580(28-29): 6537-6542. [22] Qian W, Xiao B, Wang L, et al.2018. CsINV5, a tea vacuolar invertase gene enhances cold tolerance in transgenic Arabidopsis[J]. BMC Plant Biology, 18(1): 228-248. [23] Reyna N S, Yang Y.2006. Molecular analysis of the rice MAP kinase gene family in relation to Magnaporthe grisea infection[J]. Molecular Plant-Microbe Interactions, 19(05): 530-540. [24] Rodriguez M C S, Petersen M, Mundy J.2010. Mitogen-activated protein kinase signaling in plants[J]. Plant Biology, 61(01): 621-649. [25] Teige M, Scheikl E, Eulgem T, et al.2004. The MKK2 pathway mediates cold and salt stress signaling in Arabidopsis[J]. Molecular Cell, 15(01): 141-152. [26] Vyas D, Kumar S.2005. Tea (Camellia sinensis (L.) O. Kuntze) clone with lower period of winter dormancy exhibits lesser cellular damage in response to low temperature[J]. Plant Physiology and Biochemistry, 43(04): 383-388. [27] Wang L, Cao H, Qian W, et al.2017. Identification of a novel bZIP transcription factor in Camellia sinensis as a negative regulator of freezing tolerance in transgenic Arabidopsis[J]. Annals of Botany, 119(07): 1195-1209. [28] Wang X, Qiong Y Z, Chun L M, et al.2013. Global transcriptome profiles of Camellicaoa sinensis during cold acclimation[J]. BMC Genomics, 415(14): 1471-2164. [29] Wei C L, Yang H, Wang S B, et al.2018. Draft genome sequence of Camellia sinensis var. sinensis provides insights into the evolution of the tea genome and tea quality[J]. Proceedings of the National Academy of Sciences of the USA, 115(18): E4151-E4158. [30] Wen J, Oono K, Imai R.2002. Two novel mitogen-activated protein signaling components, OsMEK1 and OsMAP1, are involved in a moderate low-temperature signaling pathway in rice[J]. Plant Physiology, 129(04): 1880-1891. [31] Xia E H, Tong W, Hou Y, et al.2020. The reference genome of tea plant and resequencing of 81 diverse accessions provide insights into its genome evolution and adaptation[J]. Molecular Plant, 13(7): 1013-1026. [32] Yamakawa H, Katou S, Seo S, et al.2004. Plant MAPK phosphatase interacts with calmodulins[J]. Journal of Biological Chemistry, 279(02): 928-936. [33] Yue C, Cao H, Wang L, et al.2015. Effects of cold acclimation on sugar metabolism and sugar-related gene expression in tea plant during the winter season[J]. Plant Molecular Biology, 88(06): 591-608. [34] Zhang Y, Zhu X, Chen X, et al.2014. Identification and characterization of cold-responsive microRNAs in tea plant (Camellia sinensis) and their targets using high-throughput sequencing and degradome analysis[J]. BMC Plant Biology, 14(01): 271-288. [35] Zhou Y, Zhang D, Pan J, et al.2012. Overexpression of a multiple stress-responsive gene, ZmMAPK4, enhances tolerance to low temperature in transgenic tobacco[J]. Plant Physiology and Biochemistry, 58: 174-181.