Research article

Transcriptome profiling in Camellia japonica var. decumbens for the discovery of genes involved in chilling tolerance under cold stress

Yawen Wu, Markus Müller, Tian Bai, Shunyang Yao, Oliver Gailing, Zhen Liu

Yawen Wu
College of Forestry, Henan Agricultural University, 450002, Zhengzhou, China
Markus Müller
University of Göttingen, Forest Genetics and Forest Tree Breeding, Büsgenweg 2, 37077 Göttingen, Germany
Tian Bai
College of Forestry, Henan Agricultural University, 450002, Zhengzhou, China
Shunyang Yao
College of Forestry, Henan Agricultural University, 450002, Zhengzhou, China
Oliver Gailing
University of Göttingen, Forest Genetics and Forest Tree Breeding, Büsgenweg 2, 37077 Göttingen, Germany
Zhen Liu
College of Forestry, Henan Agricultural University, 450002, Zhengzhou, China. Email:

Online First: June 28, 2019
Wu, Y., Müller, M., Bai, T., Yao, S., Gailing, O., Liu, Z. 2019. Transcriptome profiling in Camellia japonica var. decumbens for the discovery of genes involved in chilling tolerance under cold stress. Annals of Forest Research DOI:10.15287/afr.2018.1311

Camellia japonica var. decumbens is a naturally occurring highly cold resistant variety of Camellia japonica which is suitable for snowy and cold regions. However, the underlying cold-adaptive mechanisms associated with gene regulation have been poorly investigated. We analyzed the transcriptomic changes caused by cold stress in a cold-tolerant accession. Samples were collected at the end of each temperature treatment (T1, T3, T5, T7 and T9 represent the temperatures 25°C, 0°C, -4°C, -8°C and -12°C, respectively). Sample T1 at 25°C was used as control. Based on transcriptome analysis, 2828, 2384, 3099 and 3075 differentially expressed genes (DEGs) were up-regulated, and 3184, 2592, 2373 and 2615 DEGs were down-regulated by analyzing T3/T1, T5/T1, T7/T1 and T9/T1, respectively. A gene ontology (GO) analysis revealed an enrichment of GO terms such as response to stimulus, metabolic process, catalytic activity or binding. Out of the larger number of DEGs, 67 functional and regulatory DEGs stood out, since they were functionally characterized in other models. These genes are cold-responsive transcription factors (26) or involved in cold sensor or signal transduction (17) and in the stabilization of the plasma membrane and osmosensing response (24). These results suggest rapid and multiple molecular mechanisms of perception, transduction and responses to cold stress in cold acclimation of Camellia japonica var. decumbens. They could also serve as a valuable resource for relevant research on cold-tolerance and help to explore cold-related genes to foster the understanding of low-temperature tolerance and plant-environment interactions.

Abbasi F., Onodera H., Toki S., Tanaka H., Komatsu S., 2004. OsCDPK13, a calcium-dependent protein kinase gene from rice, is induced by cold and gibberellin in rice leaf sheath. Plant Molecular Biology 55(4):541-552. DOI: 10.1007/s11103-004-1178-y

Altschul S.F., Madden T.L., Schaffer A.A., Zhang J, Zhang Z, Miller W, et al. 1997. Gapped BLAST and PSIBLAST: a new generation of protein database search programs. Nucleic Acids Research 25(17):3389-3402. DOI: 10.1093/nar/25.17.3389

Apweiler R., Bairoch A., Wu C.H., Barker W.C., Boeckmann B., Ferro S. 2004. UniProt: the Universal Protein knowledgebase. Nucleic Acids Research 32: D115-119. DOI: 10.1093/nar/gkh131

Ashburner M., Ball C.A., Blake J.A., Botstein D., Butler H., Cherry J.M., 2000. Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nature Genetics 25(1):25-29. DOI: 10.1038/75556

Audic S., Claverie JM., 1997. The significance of digital gene expression profiles. Genome Research 7(10):986-995. DOI: 10.1101/gr.7.10.986

Chen W.J., Zhu T., (2004) Networks of transcription factors with roles in environmental stress response. Trends in Plant Scince 9(12):591-596. DOI: 10.1016/j.tplants.2004.10.007

Cheng H., Chen X., Fang J., An Z., Hu Y. and Huang. H., 2018. Comparative transcriptome analysis reveals an early gene expression profile that contributes to cold resistance in Hevea brasiliensis (the Para rubber tree). Tree Physiology 38: 1409-1423. DOI: 10.1093/treephys/tpy014

Chinnusamy V., Ohta M., Kanrar S., Lee B.H., Hong X., Agarwal M., Zhu J.K., (2003) ICE1: a regulator of cold-induced transcriptome and freezing tolerance in Arabidopsis. Genes Development 17(8): 1043-1054. DOI: 10.1101/gad.1077503

Deng Y., Li J., Wu S., Zhu Y., Chen Y., He F., 2006. Integrated nr Database in protein annotation system and its localization. Computer Engineering 32:71-76.

Dhanaraj A.L., Slovin J.P., Rowland L.J., 2004. Analysis of gene expression associated with cold acclimation in blueberry floral buds using expressed sequence tags. Plant Science 166 (4): 863-872. DOI: 10.1016/j.plantsci.2003.11.013

Dietz K. J., Vogel M. O., Viehhauser A., 2010. AP2/EREBP transcription factors are part of gene regulatory networks and integrate metabolic, hormonal and environmental signals in stress acclimation and retrograde signaling. Protoplasma 245(1-4):3-14. DOI: 10.1007/s00709-010-0142-8

Djebali S., Davis C.A., Merkel A., Dobin A., Lassmann T., Mortazavi A., 2012. Landscape of transcription in human cells. Nature 489(7414): 101-108. DOI: 10.1038/nature11233

Eddy S.R., 1998. Profile hidden Markov models. Bioinformatics 14(9): 755-763. DOI: 10.1093/bioinformatics/14.9.755

Finn R.D., Bateman A., Clements J., Coggill P., Eberhardt R. Y., Eddy S. R., 2014. Pfam: the protein families database. Nucleic Acids Research 42: D222-230. DOI: 10.1093/nar/gkt1223

Gai J., 2000. Methods of experimental statistics. Beijing: China Agriculture Press: 211-212.

Gao M.J., Schafer U.A., Parkin I.A., Hegedus D.D., Lydiate D. J., Hannoufa A., 2003. A novel protein from Brassica napus has a putative KID domain and responds to low temperature. Plant Journal 33(6): 1073-1086. DOI: 10.1046/j.1365-313X.2003.01694.x

Gaete-Loyola, J., Lagos C., Beltrán M.F., Valenzuela S., Emhart V., Fernández M., 2017. Transcriptome profiling of Eucalyptus nitens reveals deeper insight into the molecular mechanism of cold acclimation and deacclimation process. Tree Genetics and Genomics 13: 37. DOI: 10.1007/s11295-017-1121-4

Gong ., Gong Z., Guo Y., Chen X., Zhu J.K., 2002. Biochemical and functional characterization of PKS11, a novel Arabidopsis protein kinase. Journal of Biological Chemistry 277(31): 28340-28350. DOI: 10.1074/jbc.M107719200

Grabherr M.G., Haas B.J., Yassour M., Levin J.Z., Thompson D.A., Amit I., 2011. Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nature Biotechnology 29(7):644-652. DOI: 10.1038/nbt.1883

Guo Y., Sheng Q., Li J., Ye F., Samuels D.C., Shyr Y., 2013. Large scale comparison of gene expression levels by microarrays and RNAseq using TCGA data. PloS One 8(8): e71462. DOI: 10.1371/journal.pone.0071462

Hosseinpour B., Sepahvand S., Kamali Aliabad K., Bakhtiarizadeh M., Imani A., Assareh R., 2018. Transcriptome profiling of fully open flowers in a frost-tolerant almond genotype in response to freezing stress. Molecular Genetics and Genomics 293: 151-163. DOI: 10.1007/s00438-017-1371-8

Janská A., Marík P., Zelenková S., Ovesná J., 2010. Cold stress and acclimation - what is important for metabolic adjustment? Plant Biology 12(3):395-405. DOI: 10.1111/j.1438-8677.2009.00299.x

Kanehisa M., Goto S., Kawashima S., Okuno Y., Hattori M., 2004. The KEGG resource for deciphering the genome. Nucleic Acids Research 32:D277-280. DOI: 10.1093/nar/gkh063

Kim S., Jung E., Shin S., 2012. Anti-inflammatory activity of Camellia japonica oil. BMB Rep. 45:177-182. DOI: 10.5483/BMBRep.2012.45.3.177

Koonin E.V., Fedorova N.D., Jackson J.D., Jacobs A.R., Krylov D.M., Makarova K.S., 2004. A comprehensive evolutionary classification of proteins encoded in complete eukaryotic genomes. Genome Biology 5(2):R7. DOI: 10.1186/gb-2004-5-2-r7

Kondo K. 2008. Cytological studies in cultivated species of Camellia: I. diploid species and their hybrids. Japanese Journal of Breeding 27: 28-38. DOI: 10.1270/jsbbs1951.27.28

Langmead B., Trapnell C., Pop M., Salzberg S.L., 2009. Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biology. 10(3): R25. DOI: 10.1186/gb-2009-10-3-r25

Lata C., Prasad M., 2011. Role of DREBs in regulation of abiotic stress responses in plants. Journal of Experimental Botany 62(14): 4731-4748. DOI: 10.1093/jxb/err210

Leng N., Dawson J.A., Thomson J.A., Ruotti V., Rissman A.I., Smits B.M., 2013. EBSeq: an empirical Bayes hierarchical model for inference in RNA-seq experiments. Bioinformatics 29(8): 1035-1043. DOI: 10.1093/bioinformatics/btt087

Lin M.R., 1994. The trees of the Japanese. Japan, The Society For Truth And Light, pp. 482-483

Li B., Dewey C.N., 2011. RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinformatics 12:323. DOI: 10.1186/1471-2105-12-323

Li B., Takahashi D., Kawamura Y., Uemura M., 2012. Comparison of plasma membrane proteomic changes of Arabidopsis suspension-cultured cells (T87 Line) after cold and ABA treatment in association with freezing tolerance development. Plant and Cell Physiology 53(3): 543-554. DOI: 10.1093/pcp/pcs010

Li Q., Lei S., Du K., Li L., Pang X., Wang Z., 2016. RNA-seq based transcriptomic analysis uncovers α-linolenic acid and jasmonic acid biosynthesis pathways respond to cold acclimation in Camellia japonica. Scientific Reports 6: 36463. DOI: 10.1038/srep36463

Lohse M., Nagel A., Herter T., May P., Schroda M., Zrenner R., 2014. Mercator: a fast and simple web server for genome scale functional annotation of plant sequence data. Plant, Cell & Environment 37(5): 1250-1258. DOI: 10.1111/pce.12231

Min T.L., Zhang W.J., 1996. The evolution and distribution of genus Camellia. Acta Botanica Yunnanica 18 (1): 1-13.

Mortazavi A., Williams B.A., McCue K., Schaeffer L., Wold B., 2008. Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nature Methods 5(7): 621-628. DOI: 10.1038/nmeth.1226

Munnik T., 2001. Phosphatidic acid: an emerging plant lipid second messenger. Trends in Plant Science 6(5): 227-233. DOI: 10.1016/S1360-1385(01)01918-5

Piao M.J., Yoo E.S., Koh Y.S., 2011. Antioxidant effects of the ethanol extract from flower of Camellia japonica via scavenging of reactive oxygen species and induction of antioxidant enzymes. International Journal of Molecular Sciences 12: 2618-2630. DOI: 10.3390/ijms12042618

Reiner A., Yekutieli D., Benjamini Y., 2003. Identifying differentially expressed genes using false discovery rate controlling procedures. Bioinformatics 19(3): 368-375. DOI: 10.1093/bioinformatics/btf877

Renaut J., Hausman J.F., Wisniewski M., 2006. Proteomics and low temperature studies: bridging the gap between gene expression and metabolism. Physiologia Plantarum 126 (1): 97-109. DOI: 10.1111/j.1399-3054.2006.00617.x

Tatusov R.L., Galperin M.Y., Natale D.A., Koonin E.V., 2000. The COG database: a tool for genome-scale analysis of protein functions and evolution. Nucleic Acids Research 28(1): 33-36. DOI: 10.1093/nar/28.1.33

Tan H., Huang H., Tie M., 2016. Transcriptome profiling of two Asparagus Bean (Vigna unguiculata subsp. sesquipedalis) cultivars differing in chilling tolerance under cold stress. Plos One 11(3):e0151105. DOI: 10.1371/journal.pone.0151105

Thomashow M.F., 1999. Plant cold acclimation: freezing tolerance genes and regulatory mechanisms. Annual Review of Plant Physiology and Plant Molecular Biology 50: 571-599. DOI: 10.1146/annurev.arplant.50.1.571

Trapnell C., Williams B.A., Pertea G., Mortazavi A., Kwan G., van Baren M.J., 2010. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nature Biotechnology 28(5): 511-515. DOI: 10.1038/nbt.1621

Uemura M., Tominaga Y., Nakagawara C., Shigematsu S., Minami A., Kawamura Y., 2006. Responses of the plasma membrane to low temperatures. Physiology Plant 126(1): 81-89. DOI: 10.1111/j.1399-3054.2005.00594.x

Vergnolle C., Vaultier M.N., Taconnat L., Renou J.P., Kader J.C., Zachowski A., Ruelland E., 2005. The cold-induced early activation of phospholipase C and D pathways determines the response of two distinct clusters of genes in Arabidopsis cell suspensions. Plant Physiology 139: 1217-1233. DOI: 10.1104/pp.105.068171

Wang X.C., Zhao Q.Y., Ma C.L., 2013. Global transcriptome profiles of Camellia sinensis during cold acclimation. Bmc Genomics 14(1): 415. DOI: 10.1186/1471-2164-14-415

Wang H.B., Cheng L.L., Chang Y.S., 2016. Physiological and transcriptome response of apple dwarfing rootstock to cold stress and cold-resistant genes screening. Acta Horticulturae Sinica 43(8): 1437-1451

Xu Y., Chen F.., 2008. The LT50 and cold tolerance adaptability of Chrysanthemum during a natural drop in temperature. Acta Horticulturae Sinica 35(4): 559-564

Xu W., Li R., Zhang N., Ma F., Jiao Y., Wang Z., 2014. Transcriptome profiling of Vitis amurensis, an extremely cold-tolerant Chinese wild Vitis species, reveals candidate genes and events that potentially connected to cold stress. Plant Molecular Biology 86: 527-541. DOI: 10.1007/s11103-014-0245-2

Yoshikawa M., Morikawa T., Asao Y., Fujiwara E., Nakamura S., Matsuda H., 2007. Medicinal flowers. XV. The structures of noroleanane- and oleanane-type triterpene oligoglycosides with gastroprotective and platelet aggregation activities from flower buds of Camellia japonica. Chemical and Pharmaceutical Bulletin (Tokyo) 55:606-612. DOI: 10.1248/cpb.55.606

Zhao Y., Zhou Y., Jiang H., Li X., Gan D., Peng X., Zhu S., Cheng B., 2011. Systematic analysis of sequences and expression patterns of drought-responsive members of the HD-Zip gene family in maize. Plos One 6(12): e28488. DOI: 10.1371/journal.pone.0028488

Supporting Information
No metrics available for this article.

Related Articles

Related Authors


In Google Scholar

In Annals of Forest Research

In Google Scholar

  • Yawen Wu
  • Markus Müller
  • Tian Bai
  • Shunyang Yao
  • Oliver Gailing
  • Zhen Liu
  • Yawen Wu
  • Markus Müller
  • Tian Bai
  • Shunyang Yao
  • Oliver Gailing
  • Zhen Liu