Abstract
In this study, full-length (1282–1330 bp) α-expansin 1 (EXPA1) gene from three different accessions belonging to Saccharum complex (Saccharum officinarum—SoEXPA1, Erianthus arundinaceus—EaEXPA1, and Saccharum spp. hybrid—ShEXPA1) was isolated using RAGE technique and characterized. The intronic and coding regions of isolated expansin genes ranged between 526–568 and 756–762 bp, respectively. An open reading frame encoding a polypeptide of 252 amino acids was obtained from S. officinarum and commercial sugarcane hybrid, whereas 254 amino acids were obtained in E. arundinaceus, a wild relative of Saccharum. Bioinformatics analysis of deduced protein revealed the presence of specific signature sequences and conserved amino acid residues crucial for the functioning of the protein. The predicted physicochemical characterization showed that the protein is stable in nature with instability index (II) value less than 40 and also clearly shown the dominance of random coil in the protein structure. Phylogenetic analysis revealed high conservation of EXPA1 among Saccharum complex and related crop species, Sorghum bicolor and Zea mays. The docking study of EXPA1 protein showed the interaction with xylose, which is present in xyloglucan of plant cell wall, elucidated the role of the expansin proteins in plant cell wall modification. This was further supported by the subcellular localization experiment in which it is clearly seen that the expansin protein localizes in the cell wall. Relative expression analysis of EXPA1 gene in Saccharum complex during drought stress showed high expression of the EaEXPA1 in comparison with SoEXPA1 and ShEXPA1 indicating possible role of EaEXPA1 in increased water-deficit stress tolerance in E. arundinaceus. These results suggest the potential use of EXPA1 for increasing the water-deficient stress tolerance levels in crop plants.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Augustine SM, Syamaladevi DP, Premachandran MN, Ravichandran V, Subramonian N (2015) Physiological and molecular insights to drought responsiveness in Erianthus sp. Sugar Tech 17(2):121–129. https://doi.org/10.1007/s12355-014-0312-7
Azeez A, Sane AP, Tripathi SK, Bhatnagar D, Nath P (2010) The gladiolus GgEXPA1 is a GA-responsive α-expansin gene expressed ubiquitously during expansion of all floral tissues and leaves but repressed during organ senescence. Postharvest Biol Technol 58:48–56. https://doi.org/10.1016/j.postharvbio.2010.05.006
Basu A, Sarkar A (2014) Molecular docking and ligand-protein interaction study of the expansin protein AtEXPA23 and EXLX1, In: Proceedings of 1st International Science and Technology Congress, pp 1–8
Bhattacharya D, Nowotny J, Cao R, Cheng J (2016) 3Drefine: an interactive web server for efficient protein structure refinement. Nucleic Acids Res 44(W1):W406–W409. https://doi.org/10.1093/nar/gkw336
Bowie JU, Luthy R, Eisenberg D (1991) A method to identify protein sequences that fold into a known three-dimensional structure. Science 253(5016):164–170. https://doi.org/10.1126/science.1853201
Buxbaum E (2007) Fundamentals of protein structure and function, vol 31. Springer, New York. https://doi.org/10.1007/978-3-319-19920-7
Choi D, Cho HT, Lee Y (2006) Expansins: expanding importance in plant growth and development. Physiol Plant 126(4):511–518. https://doi.org/10.1111/j.1399-3054.2006.00612.x
Chomczynski P, Mackey K (1995) Modification of the TRI reagent DNA/protein isolation procedure for isolation of RNA from polysaccharide and proteoglycan rich sources. Biotechniques 19:942–945
Chujo T, Takai R, Akimoto-Tomiyama C, Ando S, Minami E, Nagamura Y, Kaku H, Shibuya N, Yasuda M, Nakashita H, Umemura K (2007) Involvement of the elicitor-induced gene OsWRKY53 in the expression of defense-related genes in rice. Biochim Biophys Acta Gene Struct Expr 1769(7–8):497–505. https://doi.org/10.1016/j.bbaexp.2007.04.006
Cosgrove DJ (2000a) Loosening of plant cell walls by expansins. Nature 407:321–326. https://doi.org/10.1038/35030000
Cosgrove DJ (2000b) New genes and new biological roles for expansins. Curr Opin Plant Biol 3(1):73–78. https://doi.org/10.1016/S1369-5266(99)00039-4
Cosgrove DJ (2015) Plant expansins: diversity and interactions with plant cell walls. Curr Opin Plant Biol 25:162–172. https://doi.org/10.1016/j.pbi.2015.05.014
Dharshini S, Chakravarthi M, Ashwin Narayan J, Manoj VM, Naveenarani M, Kumar R, Meena M, Ram B, Appunu C (2016) De novo sequencing and transcriptome analysis of a low temperature tolerant Saccharum spontaneum clone IND 00-1037. J Biotechnol 231:280–294. https://doi.org/10.1016/j.jbiotec.2016.05.036
Eisenberg D, Luthy R, Bowie JU (1997) VERIFY3D: assessment of protein models with three-dimensional profiles. Methods Enzymol 277:396–404. https://doi.org/10.1038/356083a0
Garnier J, Gibrat JF, Robson B (1996) GOR secondary structure prediction method version IV. Methods Enzymol 266:540–553
Gasteiger E, Hoogland C, Gattiker A, Duvaud S, Wilkins, Appel RD, Bairoch A (2005) Protein identification and analysis tools on the ExPASy server. In: Walker JM (ed) The proteomics protocols handbook. Human Press, New York, pp 571–607. https://doi.org/10.1385/1-59259-890-0:571
Grosdidier A, Zoete V, Michielin O (2011) SwissDock, a protein-small molecule docking web service based on EADock DSS. Nucleic Acids Res 39(2):270–277. https://doi.org/10.1093/nar/gkr366
Guermeur Y (1997) Combinaison de classifieurs statistiques, Application a la prediction de structure secondaire des proteins. Ph. D. Thesis, Universite Paris
Guruprasad K, Reddy BVP, Pandit MW (1990) Correlation between stability of a protein and its dipeptide composition: a novel approach for predicting in vivo stability of a protein from its primary sequence. Protein Eng Des Sel 4:155–164. https://doi.org/10.1093/protein/4.2.155
Harmer S, Orford S, Timmis J (2002) Characterisation of six α-expansin genes in Gossypium hirsutum (upland cotton). Mol Genet Genom 268(1):1–9. https://doi.org/10.1007/s00438-002-0721-2
Haseloff J, Siemering KR (1998) The uses of green fluorescent protein in plants. In: Chalfie M, Kain SR (eds) Green fluorescent protein: properties, applications, and protocols. Wiley, Chichester, pp 191–220
Ikai AJ (1980) Thermo stability and aliphatic index of globular proteins. J Biochem 88:1895–1898. https://doi.org/10.1093/oxfordjournals.jbchem.a133168
Jones L, McQueen-Mason S (2004) A role for expansins in dehydration and rehydration of the resurrection plant Craterostigma plantagineum. FEBS Lett 559(1–3):61–65. https://doi.org/10.1016/S0014-5793(04)00023-7
Jung J, O’Donoghue EM, Dijkwel PP, Brummell DA (2010) Expression of multiple expansin genes is associated with cell expansion in potato organs. Plant Sci 179:77–85. https://doi.org/10.1016/j.plantsci.2010.04.007
Kallberg M, Wang H, Wang S, Peng J, Wang Z, Lu H, Xu J (2012) Template-based protein structure modeling using the RaptorX web server. Nat Protoc 7(8):1511–1522. https://doi.org/10.1038/nprot.2012.085
Kuluev B, Avalbaev A, Mikhaylova E, Nikonorov Y, Berezhneva Z, Chemeris A (2016) Expression profiles and hormonal regulation of tobacco expansin genes and their involvement in abiotic stress response. J Plant Physiol 206:1–12. https://doi.org/10.1016/j.jplph.2016.09.001
Kyte J, Doolottle RF (1982) A simple method for displaying the hydropathic character of a protein. J Mol Biol 157(1):105–132. https://doi.org/10.1016/0022-2836(82)90515-0
Lakhanpal N, Verma D, Kaur R, Singh K (2018) Characterization of cold responsive uncoupling protein1 (UCP1) gene from Brassica juncea L. (Czern. and Coss.). J Plant Biochem Biotechnol 27(1):108–117. https://doi.org/10.1007/s13562-017-0421-y
Lin C, Choi HS, Cho HT (2011) Root hair-specific EXPANSIN A7 is required for root hair elongation in Arabidopsis. Mol Cells 31(4):393–397. https://doi.org/10.1007/s10059-011-0046-2
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using realtime quantitative PCR and the 2−ΔΔCT method. Methods 25(4):402–408. https://doi.org/10.1006/meth.2001.1262
Lovell SC, Davis IW, Arendall WB III, de Bakker PI, Word JM, Prisant MG, Richardson JS, Richardson DC (2003) Structure validation by Calpha geometry: phi, psi and Cbeta deviation. Proteins: structure. Funct Genet 50:437–450. https://doi.org/10.1002/prot.10286
Lu P, Kang M, Jiang X, Dai F, Gao J, Zhang C (2013) RhEXPA4, a rose expansin gene, modulates leaf growth and confers drought and salt tolerance to Arabidopsis. Planta 237(6):1547–1559. https://doi.org/10.1007/s00425-013-1867-3
Luthy R, Bowie JU, Eisenberg D (1992) Assessment of protein models with three-dimensional profiles. Nature 356(6364):83–85. https://doi.org/10.1038/356083a0
Marowa P, Ding A, Kong Y (2016) Expansins: roles in plant growth and potential applications in crop improvement. Plant Cell Rep 35(5):949–965. https://doi.org/10.1007/s00299-016-1948-4
McGuffin LJ, Bryson K, Jones DT (2000) The PSIPRED protein structure prediction server. Bioinformatics 16(4):404–405. https://doi.org/10.1093/bioinformatics/16.4.404
Mukherjee SK (1957) Origin and distribution of Saccharum. Bot Gaz 119:55–61. https://doi.org/10.1086/335962
Park JW, Benatti TR, Marconi T, Yu Q, Solis-Gracia N, Mora V, da Silva JA (2015) Cold responsive gene expression profiling of sugarcane and Saccharum spontaneum with functional analysis of a cold inducible saccharum homolog of NOD26-like intrinsic protein to salt and water stress. PLoS One 10(5):e0125810. https://doi.org/10.1371/journal.pone.0125810
Petersen TN, Brunak S, von Heijne G, Nielsen H (2011) SignalP 4.0: discriminating signal peptides from trans-membrane regions. Nat Methods 8(10):785–786. https://doi.org/10.1038/nmeth.1701
Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, Ferrin TE (2004) UCSF Chimera—a visualization system for exploratory research and analysis. J Comput Chem 25(13):1605–1612. https://doi.org/10.1002/jcc.20084
Sampedro J, Cosgrove DJ et al (2005a) The expansin superfamily. Genome Biol 6(12):242. https://doi.org/10.1186/gb-2005-6-12-242
Sampedro J, Lee Y, Carey RE, dePamphilis C, Cosgrove DJ (2005b) Use of genomic history to improve phylogeny and understanding of births and deaths in a gene family. Plant J 44(3):409–419. https://doi.org/10.1111/j.1365-313X.2005.02540.x
Santiago TR, Pereira VM, de Souza WR, Steindorff AS, Cunha BA, Gaspar M, Fávaro LC, Formighieri EF, Kobayashi AK, Molinari HB (2018) Genome-wide identification, characterization and expression profile analysis of expansins gene family in sugarcane (Saccharum spp.). PloS One 13(1):e0191081. https://doi.org/10.1371/journal.pone.0191081
Selvarajan D, Mohan C, Dhandapani V, Nerkar G, Jayanarayanan AN, Mohanan MV, Murugan N, Kaur L, Chennappa M, Kumar R, Meena M (2018) Differential gene expression profiling through transcriptome approach of Saccharum spontaneum L. under low temperature stress reveals genes potentially involved in cold acclimation. 3 Biotech 8(4):195. https://doi.org/10.1007/s13205-018-1194-2
Stewart C (2001) The utility of green fluorescent protein in transgenic plants. Plant Cell Rep 20(5):376–382. https://doi.org/10.1007/s002990100346
Sun T, Zhang Y, Chai T (2011) Cloning, characterization, and expression of the BjEXPA1 gene and its promoter region from Brassica juncea L. Plant Growth Regul 64(1):39–51. https://doi.org/10.1007/s10725-010-9533-2
Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30:2725–2729. https://doi.org/10.1093/molbev/mst197
Tan J, Wang M, Shi Z, Miao X (2018) OsEXPA10 mediates the balance between growth and resistance to biotic stress in rice. Plant Cell Rep 37:993–1002. https://doi.org/10.1007/s00299-018-2284-7
Vannerum K, Huysman MJ, De Rycke R, Vuylsteke M, Leliaert F, Pollier J, Lutz-Meindl U, Gillard J, De Veylder L, Goossens A, Inze D (2011) Transcriptional analysis of cell growth and morphogenesis in the unicellular green alga Micrasterias (Streptophyta), with emphasis on the role of expansin. BMC Plant Biol 11(1):128. https://doi.org/10.1186/1471-2229-11-128
Wu YJ, Cosgrove DJ (2000) Adaptation of roots to low water potentials by changes in cell wall extensibility and cell wall proteins. J Exp Bot 51(350):1543–1553. https://doi.org/10.1093/jexbot/51.350.1543
Wu Y, Thorne ET, Sharp RE, Cosgrove DJ (2001) Modification of expansin transcript levels in the maize primary root at low water potentials. Plant Physiol 126(4):1471–1479. https://doi.org/10.1104/pp.126.4.1471
Xu J, Tian J, Belanger FC, Huang B (2007) Identification and characterization of an expansin gene AsEXP1 associated with heat tolerance in C3 Agrostis grass species. J Exp Bot 58(13):3789–3796. https://doi.org/10.1093/jxb/erm229
Xu Q, Xu X, Shi Y, Xu J, Huang B (2014) Transgenic tobacco plants overexpressing a grass PpEXP1 gene exhibit enhanced tolerance to heat stress. PLoS One 9(7):e100792. https://doi.org/10.1371/journal.pone.0100792
Yang L, Zheng B, Mao C, Qi X, Liu F, Wu P (2004) Analysis of transcripts that are differentially expressed in three sectors of the rice root system under water deficit. Mol Genet Genom 272(4):433–442. https://doi.org/10.1007/s00438-004-1066-9
Zenoni S, Fasoli M, Tornielli GB, Dal Santo S, Sanson A, de Groot P, Sordo S, Citterio S, Monti F, Pezzotti M (2011) Overexpression of PhEXPA1 increases cell size, modifies cell wall polymer composition and affects the timing of axillary meristem development in Petunia hybrida. New Phytol 191(3):662–677. https://doi.org/10.1111/j.1469-8137.2011.03726.x
Acknowledgements
The authors thank the Director, ICAR-Sugarcane Breeding Institute, Coimbatore, Tamil Nadu, India, for providing the facilities. This work was supported by the Department of Biotechnology (DBT) (Grant no. 102/IFD/SAN/1151/2017-2018), Government of India. One of the authors, Ashwin Narayan J, thanks Council of Scientific and Industrial Research (CSIR), New Delhi, India, for the award of Senior Research Fellowship (SRF) (09/706/0003/2018-EMR-I).
Author information
Authors and Affiliations
Contributions
JAN, NS, MNP and CA conceived and designed the research. JAN, SD, VMN, TSSP, KK and GSS performed the experiments. JAN, MNP and CA analyzed the results and drafted the manuscript. BR revised the manuscript.
Corresponding authors
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Narayan, J.A., Dharshini, S., Manoj, V.M. et al. Isolation and characterization of water-deficit stress-responsive α-expansin 1 (EXPA1) gene from Saccharum complex. 3 Biotech 9, 186 (2019). https://doi.org/10.1007/s13205-019-1719-3
Received:
Accepted:
Published:
DOI: https://doi.org/10.1007/s13205-019-1719-3