Advertisement

Functional & Integrative Genomics

, Volume 18, Issue 5, pp 489–503 | Cite as

Transcriptome-wide identification and expression profile analysis of the bHLH family genes in Camellia sinensis

  • Xin Cui
  • Yong-Xin Wang
  • Zhi-Wei Liu
  • Wen-Li Wang
  • Hui Li
  • Jing Zhuang
Original Article

Abstract

The tea plant is an important commercial horticulture crop cultivated worldwide. Yield and quality of this plant are influenced by abiotic stress. The bHLH family transcription factors play a pivotal role in the growth and development, including abiotic stress response, of plants. A growing number of bHLH proteins have been functionally characterized in plants. However, few studies have focused on the bHLH proteins in tea plants. In this study, 120 CsbHLH TFs were identified from tea plants using computational prediction method. Structural analysis detected 23 conservative residues, with over 50% identities in the bHLH domain. Moreover, 103 CsbHLH proteins were assumed to bind DNA and encompassed 98 E-Box binders and 85 G-Box binders. The CsbHLH proteins were grouped into 20 subfamilies based on phylogenetic analysis and a previous classification system. A survey of transcriptome profiling screened 22 and 39 CsbHLH genes that were upregulated under heat and drought stress. Nine CsbHLH genes were validated using qRT-PCR. Results were approximately in accordance with transcriptome data. These genes could be induced by one or more abiotic stresses.

Keywords

bHLH Transcription factor Phylogenetic analysis Expression pattern Abiotic stress Tea plant 

Abbreviations:

ABA

Abscisic acid

AIB

ABA-inducible bHLH-type transcription factor

bHLH

Basic helix-loop-helix

BLAST

Basic Local Alignment Search Tool

BRs

Brassinosteroids

CBF

C-repeat binding factor

COE

Collier/Olf-1/EBF

FPKM

Fragments per kilobase of transcript per million mapped reads

GO

Gene ontology

HMM

Hidden Markov model

JA

Jasmonic acid

MEGA

Molecular evolutionary genetics analysis

MEME

Multiple Em for motif elicitation

NCBI

National Center for Biotechnology Information

NJ

Neighbor-joining

PAS

Per-Arnt-Sim

PEG

Polyethylene glycol

qRT-PCR

Quantitative real-time polymerase chain reaction

RNA-Seq

RNA sequencing

TF

Transcription factor

Notes

Acknowledgements

The research was supported by the National Natural Science Foundation of China (31570691).

Author’s contributions

Conceived and designed the experiments: JZ, XC. Performed the experiments: XC, YXW, ZWL, WLW, HL. Analyzed the data: XC, YXW, JZ. Contributed reagents/materials/analysis tools: JZ. Wrote the paper: XC. Revised the paper: JZ, XC, YXW. All authors read and approved the final manuscript.

Supplementary material

10142_2018_608_Fig11_ESM.gif (369 kb)
Figure S1

Multiple alignments of CsbHLH proteins (GIF 368 kb)

10142_2018_608_MOESM1_ESM.tif (10.3 mb)
High resolution image (TIFF 10597 kb)
10142_2018_608_MOESM2_ESM.xlsx (37 kb)
Table S1 (XLSX 37 kb)
10142_2018_608_MOESM3_ESM.xlsx (122 kb)
Table S2 (XLSX 121 kb)
10142_2018_608_MOESM4_ESM.xlsx (19 kb)
Table S3 (XLSX 18 kb)
10142_2018_608_MOESM5_ESM.xlsx (21 kb)
Table S4 (XLSX 20 kb)
10142_2018_608_MOESM6_ESM.xlsx (39 kb)
Table S5 (XLSX 38 kb)
10142_2018_608_MOESM7_ESM.xlsx (11 kb)
Table S6 (XLSX 10 kb)
10142_2018_608_MOESM8_ESM.xlsx (37 kb)
Table S7 (XLSX 36 kb)

References

  1. Abe H, Yamaguchi-Shinozaki K, Urao T, Iwasaki T, Hosokawa D, Shinozaki K (1997) Role of Arabidopsis MYC and MYB homologs in drought-and abscisic acid-regulated gene expression. Plant Cell 9(10):1859–1868PubMedPubMedCentralGoogle Scholar
  2. Ahmad A, Niwa Y, Goto S, Ogawa T, Shimizu M, Suzuki A, Kobayashi K, Kobayashi H (2015) bHLH106 integrates functions of multiple genes through their G-box to confer salt tolerance on Arabidopsis. PLoS One 10(5):e0126872CrossRefPubMedPubMedCentralGoogle Scholar
  3. Akula R, Ravishankar GA (2011) Influence of abiotic stress signals on secondary metabolites in plants. Plant Signal Behav 6(11):1720–1731CrossRefGoogle Scholar
  4. Atchley WR, Fitch WM (1997) A natural classification of the basic helix–loop–helix class of transcription factors. Proc Natl Acad Sci U S A 94(10):5172–5176CrossRefPubMedPubMedCentralGoogle Scholar
  5. Atchley WR, Terhalle W, Dress A (1999) Positional dependence, cliques, and predictive motifs in the bHLH protein domain. J Mol Evol 48(5):501–516CrossRefPubMedGoogle Scholar
  6. Bailey PC, Martin C, Toledo-Ortiz G, Quail PH, Huq E, Heim MA, Jakoby M, Werber M, Weisshaar B (2003) Update on the basic helix-loop-helix transcription factor gene family in Arabidopsis thaliana. Plant Cell 15(5):2497–2502CrossRefPubMedPubMedCentralGoogle Scholar
  7. Capella-Gutiérrez S, Silla-Martínez JM, Gabaldón T (2009) trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 25(15):1972–1973CrossRefPubMedPubMedCentralGoogle Scholar
  8. Carretero-Paulet L, Galstyan A, Roig-Villanova I, Martínez-García JF, Bilbao-Castro JR, Robertson DL (2010) Genome-wide classification and evolutionary analysis of the bHLH family of transcription factors in Arabidopsis, poplar, rice, moss, and algae. Plant Physiol 153(3):1398–1412CrossRefPubMedPubMedCentralGoogle Scholar
  9. Castillon A, Shen H, Huq E (2007) Phytochrome interacting factors: central players in phytochrome-mediated light signaling networks. Trends Plant Sci 12(11):514–521CrossRefPubMedGoogle Scholar
  10. Chen Y, Yu M, Xu J, Chen X, Shi J (2009) Differentiation of eight tea (Camellia sinensis) cultivars in China by elemental fingerprint of their leaves. J Sci Food Agric 89(14):2350–2355CrossRefGoogle Scholar
  11. 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 Dev 17(8):1043–1054CrossRefPubMedPubMedCentralGoogle Scholar
  12. Conesa A, Götz S, García-Gómez JM, Terol J, Talón M, Robles M (2005) Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 21(18):3674–3676CrossRefPubMedGoogle Scholar
  13. Deng W, Wang Y, Liu Z, Cheng H, Xue Y (2014) HemI: a toolkit for illustrating heatmaps. PLoS One 9(11):e111988CrossRefPubMedPubMedCentralGoogle Scholar
  14. Duek PD, Fankhauser C (2003) HFR1, a putative bHLH transcription factor, mediates both phytochrome a and cryptochrome signalling. Plant J 34(6):827–836CrossRefPubMedGoogle Scholar
  15. Eddy SR (2008) A probabilistic model of local sequence alignment that simplifies statistical significance estimation. PLoS Comput Biol 4(5):e1000069CrossRefPubMedPubMedCentralGoogle Scholar
  16. Feller A, Machemer K, Braun EL, Grotewold E (2011) Evolutionary and comparative analysis of MYB and bHLH plant transcription factors. Plant J 66(1):94–116CrossRefPubMedGoogle Scholar
  17. Fischer S, Brunk BP, Chen F, Gao X, Harb OS, Iodice JB, Shanmugam D, Roos DS, Stoeckert CJ (2011) Using OrthoMCL to assign proteins to OrthoMCL-DB groups or to cluster proteomes into new Ortholog groups. Curr Protoc Bioinformatics 6(12):11–16 12. 19Google Scholar
  18. Friedrichsen DM, Nemhauser J, Muramitsu T, Maloof JN, Alonso J, Ecker JR, Furuya M, Chory J (2002) Three redundant brassinosteroid early response genes encode putative bHLH transcription factors required for normal growth. Genetics 162(3):1445–1456PubMedPubMedCentralGoogle Scholar
  19. Fursova OV, Pogorelko GV, Tarasov VA (2009) Identification of ICE2, a gene involved in cold acclimation which determines freezing tolerance in Arabidopsis thaliana. Gene 429(1–2):98–103CrossRefPubMedGoogle Scholar
  20. Gilmour SJ, Sebolt AM, Salazar MP, Everard JD, Thomashow MF (2000) Overexpression of the Arabidopsis CBF3transcriptional activator mimics multiple biochemical changes associated with cold acclimation. Plant Physiol 124(4):1854–1865CrossRefPubMedPubMedCentralGoogle Scholar
  21. Goodrich J, Carpenter R, Coen ES (1992) A common gene regulates pigmentation pattern in diverse plant species. Cell 68(5):955–964CrossRefPubMedGoogle Scholar
  22. Gremski K, Ditta G, Yanofsky MF (2007) The HECATE genes regulate female reproductive tract development in Arabidopsis thaliana. Development 134(20):3593–3601CrossRefPubMedGoogle Scholar
  23. Haak DC, Fukao T, Grene R, Hua Z, Ivanov R, Perrella G, Li S (2017) Multilevel regulation of abiotic stress responses in plants. Front Plant Sci 8:1564CrossRefPubMedPubMedCentralGoogle Scholar
  24. Hall T (2011) BioEdit: an important software for molecular biology. GERF bull. Biosci 2(1):60–61Google Scholar
  25. Heim MA, Jakoby M, Werber M, Martin C, Weisshaar B, Bailey PC (2003) The basic helix-loop-helix transcription factor family in plants: a genome-wide study of protein structure and functional diversity. Mol Biol Evol 20(5):735–747.  https://doi.org/10.1093/molbev/msg088 CrossRefPubMedGoogle Scholar
  26. Jiang Y, Yang B, Deyholos MK (2009) Functional characterization of the Arabidopsis bHLH92 transcription factor in abiotic stress. Mol Gen Genomics 282(5):503–516CrossRefGoogle Scholar
  27. Jin J, Tian F, Yang D-C, Meng Y-Q, Kong L, Luo J, Gao G (2017) PlantTFDB 4.0: toward a central hub for transcription factors and regulatory interactions in plants. Nucleic Acids Res 45(D1):D1040–D1045CrossRefPubMedGoogle Scholar
  28. Kavas M, Baloğlu MC, Atabay ES, Ziplar UT, Daşgan HY, Ünver T (2016) Genome-wide characterization and expression analysis of common bean bHLH transcription factors in response to excess salt concentration. Mol Gen Genomics 291(1):129–143CrossRefGoogle Scholar
  29. Kiribuchi K, Jikumaru Y, Kaku H, MINAMI E, HASEGAWA M, KODAMA O, SETO H, OKADA K, NOJIRI H, YAMANE H (2005) Involvement of the basic helix-loop-helix transcription factor RERJ1 in wounding and drought stress responses in rice plants. Biosci Biotechnol Biochem 69(5):1042–1044CrossRefPubMedGoogle Scholar
  30. Komatsu M, Maekawa M, Shimamoto K, Kyozuka J (2001) The LAX1 and FRIZZY PANICLE 2 genes determine the inflorescence architecture of rice by controlling rachis-branch and spikelet development. Dev Biol 231(2):364–373CrossRefPubMedGoogle Scholar
  31. Kondou Y, Nakazawa M, Kawashima M, Ichikawa T, Yoshizumi T, Suzuki K, Ishikawa A, Koshi T, Matsui R, Muto S (2008) RETARDED GROWTH OF EMBRYO1, a new basic helix-loop-helix protein, expresses in endosperm to control EMBRYO growth. Plant Physiol 147(4):1924–1935CrossRefPubMedPubMedCentralGoogle Scholar
  32. Kurbidaeva A, Novokreshchenova M, Ezhova T (2015) ICE genes in Arabidopsis thaliana: clinal variation in DNA polymorphism and sequence diversification. Biol Plant 59:245–252CrossRefGoogle Scholar
  33. Larkin MA, Blackshields G, Brown N, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R (2007) Clustal W and Clustal X version 2.0. Bioinformatics 23(21):2947–2948CrossRefPubMedGoogle Scholar
  34. Ledent V, Vervoort M (2001) The basic helix-loop-helix protein family: comparative genomics and phylogenetic analysis. Genome Res 11(5):754–770CrossRefPubMedPubMedCentralGoogle Scholar
  35. Lee C-M, Thomashow MF (2012) Photoperiodic regulation of the C-repeat binding factor (CBF) cold acclimation pathway and freezing tolerance in Arabidopsis thaliana. Proc Natl Acad Sci U S A 109(37):15054–15059CrossRefPubMedPubMedCentralGoogle Scholar
  36. Lee B-H, Henderson DA, Zhu J-K (2005) The Arabidopsis cold-responsive transcriptome and its regulation by ICE1. Plant Cell 17(11):3155–3175CrossRefPubMedPubMedCentralGoogle Scholar
  37. Li X, Duan X, Jiang H, Sun Y, Tang Y, Yuan Z, Guo J, Liang W, Chen L, Yin J (2006) Genome-wide analysis of basic/helix-loop-helix transcription factor family in rice and Arabidopsis. Plant Physiol 141(4):1167–1184CrossRefPubMedPubMedCentralGoogle Scholar
  38. Li H, Sun J, Xu Y, Jiang H, Wu X, Li C (2007) The bHLH-type transcription factor AtAIB positively regulates ABA response in Arabidopsis. Plant Mol Biol 65(5):655–665CrossRefPubMedGoogle Scholar
  39. Li MY, Wang F, Jiang Q, Ma J, Xiong AS (2014) Identification of SSRs and differentially expressed genes in two cultivars of celery (Apium graveolens L.) by deep transcriptome sequencing. Hortic Res 1:10CrossRefPubMedPubMedCentralGoogle Scholar
  40. Liljegren SJ, Roeder AH, Kempin SA, Gremski K, Østergaard L, Guimil S, Reyes DK, Yanofsky MF (2004) Control of fruit patterning in Arabidopsis by INDEHISCENT. Cell 116(6):843–853CrossRefPubMedGoogle Scholar
  41. Lindemose S, O'Shea C, Jensen MK, Skriver K (2013) Structure, function and networks of transcription factors involved in abiotic stress responses. Int J Mol Sci 14(3):5842–5878CrossRefPubMedPubMedCentralGoogle Scholar
  42. Liu W, Tai H, Li S, Gao W, Zhao M, Xie C, Li WX (2014) bHLH122 is important for drought and osmotic stress resistance in Arabidopsis and in the repression of ABA catabolism. New Phytol 201(4):1192–1204CrossRefPubMedGoogle Scholar
  43. Liu Z-W, Wu Z-J, Li X-H, Huang Y, Li H, Wang Y-X, Zhuang J (2016) Identification, classification, and expression profiles of heat shock transcription factors in tea plant (Camellia sinensis) under temperature stress. Gene 576:52–59CrossRefPubMedGoogle Scholar
  44. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods 25(4):402–408CrossRefPubMedGoogle Scholar
  45. Lou W, Sun S, Wu L, Sun K (2015) Effects of climate change on the economic output of the Longjing-43 tea tree. 1972–2013. Int J Biometeorol 59(5):593–603CrossRefPubMedGoogle Scholar
  46. Ludwig SR, Habera LF, Dellaporta SL, Wessler SR (1989) Lc, a member of the maize R gene family responsible for tissue-specific anthocyanin production, encodes a protein similar to transcriptional activators and contains the myc-homology region. Proc Natl Acad Sci U S A 86(18):7092–7096CrossRefPubMedPubMedCentralGoogle Scholar
  47. Murre C, McCaw PS, Baltimore D (1989) A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell 56(5):777–783CrossRefPubMedGoogle Scholar
  48. Nesi N, Debeaujon I, Jond C, Pelletier G, Caboche M, Lepiniec L (2000) The TT8 gene encodes a basic helix-loop-helix domain protein required for expression of DFR and BAN genes in Arabidopsis siliques. Plant Cell 12(10):1863–1878CrossRefPubMedPubMedCentralGoogle Scholar
  49. Pires N, Dolan L (2010) Origin and diversification of basic-helix-loop-helix proteins in plants. Mol Biol Evol 27(4):862–874CrossRefPubMedGoogle Scholar
  50. Quattrocchio F, Wing JF, Leppen HT, Mol JN, Koes RE (1993) Regulatory genes controlling anthocyanin pigmentation are functionally conserved among plant species and have distinct sets of target genes. Plant Cell 5(11):1497–1512CrossRefPubMedPubMedCentralGoogle Scholar
  51. Rajani S, Sundaresan V (2001) The Arabidopsis myc/bHLH gene ALCATRAZ enables cell separation in fruit dehiscence. Curr Biol 11(24):1914–1922CrossRefPubMedGoogle Scholar
  52. Riechmann JL, Ratcliffe OJ (2000) A genomic perspective on plant transcription factors. Curr Opin Plant Biol 3(5):423–434CrossRefPubMedGoogle Scholar
  53. Sailsbery JK, Dean RA (2012) Accurate discrimination of bHLH domains in plants, animals, and fungi using biologically meaningful sites. BMC Evol Biol 12:154CrossRefPubMedPubMedCentralGoogle Scholar
  54. Sakamoto W, Ohmori T, Kageyama K, Miyazaki C, Saito A, Murata M, Noda K, Maekawa M (2001) The purple leaf (Pl) locus of rice: the Plw allele has a complex organization and includes two genes encoding basic helix-loop-helix proteins involved in anthocyanin biosynthesis. Plant Cell Physiol 42(9):982–991CrossRefPubMedGoogle Scholar
  55. Song XM, Huang ZN, Duan WK, Ren J, Liu TK, Li Y, Hou XL (2014) Genome-wide analysis of the bHLH transcription factor family in Chinese cabbage (Brassica rapa ssp. pekinensis). Mol Gen Genomics 289(1):77–91CrossRefGoogle Scholar
  56. Sun H, Fan HJ, Ling HQ (2015) Genome-wide identification and characterization of the bHLH gene family in tomato. BMC Genomics 16:9CrossRefPubMedPubMedCentralGoogle Scholar
  57. Szécsi J, Joly C, Bordji K, Varaud E, Cock JM, Dumas C, Bendahmane M (2006) BIGPETALp, a bHLH transcription factor is involved in the control of Arabidopsis petal size. T. EMBO J 25(16):3912–3920CrossRefPubMedPubMedCentralGoogle Scholar
  58. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: Molecular Evolutionary Genetics Analysis Version 6.0. Mol Biol Evol 30(12):2725–2729CrossRefPubMedPubMedCentralGoogle Scholar
  59. Toledo-Ortiz G, Huq E, Quail PH (2003) The Arabidopsis basic/helix-loop-helix transcription factor family. Plant Cell 15(8):1749–1770CrossRefPubMedPubMedCentralGoogle Scholar
  60. Wang Y, Jiang C-J, Li Y-Y, Wei C-L, Deng W-W (2012) CsICE1 and CsCBF1: two transcription factors involved in cold responses in Camellia sinensis. Plant Cell Rep 31(1):27–34CrossRefPubMedGoogle Scholar
  61. Wang Y-X, Liu Z-W, Wu Z-J, Li H, Zhuang J (2016a) Transcriptome-wide identification and expression analysis of the NAC gene family in tea plant [Camellia sinensis (L.) O. Kuntze]. PLoS One 11(11):e0166727CrossRefPubMedPubMedCentralGoogle Scholar
  62. Wang W, Xin H, Mingle Wang QM, Wang L, Kaleri NA, Wang Y, Li X (2016b) Transcriptomic analysis reveals the molecular mechanisms of drought-stress-induced decreases in Camellia sinensis leaf quality. Front Plant Sci 7:385PubMedPubMedCentralGoogle Scholar
  63. Wu Z-J, Li XH, Liu ZW, Xu Z-S, Zhuang J (2014) De novo assembly and transcriptome characterization: novel insights into catechins biosynthesis in Camellia sinensis. BMC Plant Biol 14:277CrossRefPubMedPubMedCentralGoogle Scholar
  64. Yin J, Chang X, Kasuga T, Bui M, Reid MS, Jiang CZ (2015) A basic helix-loop-helixtranscription factor, PhFBH4, regulates flower senescence by modulating ethylene biosynthesis pathway in petunia. Hortic Res 2:15059CrossRefPubMedPubMedCentralGoogle Scholar
  65. Yue H, Wang M, Liu S, Du X, Song W, Nie X (2016) Transcriptome-wide identification and expression profiles of the WRKY transcription factor family in broomcorn millet (Panicum miliaceum L.) BMC Genomics 17:343CrossRefPubMedPubMedCentralGoogle Scholar
  66. Zhang W, Sun Y, Timofejeva L, Chen C, Grossniklaus U, Ma H (2006) Regulation of Arabidopsis tapetum development and function by DYSFUNCTIONAL TAPETUM1 (DYT1) encoding a putative bHLH transcription factor. Development 133(16):3085–3095CrossRefPubMedGoogle Scholar
  67. Zhao L, Gao L, Wang H, Chen X, Wang Y, Yang H, Wei C, Wan X, Xia T (2013) The R2R3-MYB, bHLH, WD40, and related transcription factors in flavonoid biosynthesis. Funct Integr Genomics 13(1):75–98CrossRefPubMedGoogle Scholar
  68. Zhou L, Xu H, Mischke S, Meinhardt LW, Zhang D, Zhu X, Li X, Fang W (2014) Exogenous abscisic acid significantly affects proteome in tea plant (Camellia sinensis) exposed to drought stress. Hortic Res 1:14029CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Xin Cui
    • 1
  • Yong-Xin Wang
    • 1
  • Zhi-Wei Liu
    • 1
  • Wen-Li Wang
    • 1
  • Hui Li
    • 1
  • Jing Zhuang
    • 1
  1. 1.Tea Science Research Institute, College of HorticultureNanjing Agricultural UniversityNanjingChina

Personalised recommendations