Plant Cell Reports

, Volume 36, Issue 1, pp 179–191 | Cite as

Heterologous expression of a novel Zoysia japonica salt-induced glycine-rich RNA-binding protein gene, ZjGRP, caused salt sensitivity in Arabidopsis

  • Ke Teng
  • Penghui Tan
  • Guozeng Xiao
  • Liebao Han
  • Zhihui ChangEmail author
  • Yuehui ChaoEmail author
Original Article


Key message

A novel Zoysia japonica salt-induced glycine-rich RNA-binding protein gene was cloned in this study and its overexpression caused salt sensitivity in transgenic Arabidopsis.


Glycine-rich RNA-binding proteins (GRPs) play crucial roles in diverse plant developmental processes. However, the mechanisms and functions of GRPs in salinity stress responses remain largely unknown. In this study, rapid amplification of cDNA end (RACE) PCR methods was adopted to isolate ZjGRP from Zosyia japonica, a salt-tolerant grass species. ZjGRP cDNA was 456 bp in length, corresponding to 151 amino acids. ZjGRP was localized in the nucleus and cytoplasm, and was found particularly abundantly in stomatal guard cells. Quantitative real-time PCR showed that ZjGRP was expressed in the roots, stems, and leaves of Zoysia japonica, with the greatest expression seen in the fast-growing leaves. Furthermore, expression of ZjGRP was strongly induced by treatment with NaCl, ABA, MeJA, and SA. Overexpression of ZjGRP in Arabidopsis reduced the rate of germination and retarded seedling growth. ZjGRP-overexpressing Arabidopsis thaliana exhibited weakened salinity tolerance, likely as a result of effects on ion transportation, osmosis, and antioxidation. This study indicates that ZjGRP plays an essential role in inducing salt sensitivity in transgenic plants.


Zoysia japonica GRPs Salinity stress Transgenic Arabidopsis Gene expression 



RNA-binding proteins


Quantitative real-time polymerase chain reactions


Abscisic acid


Methyl jasmonate


Salicylic acid






Reactive oxygen species


Superoxide dismutase




Ascorbate peroxidase



This research was supported by the National High Technology Research and Development Program of China (863 Program) (No. 2013AA102607), Knowledge Innovation Program of Shen Zhen (No.JCYJ20160331151245672) and National Natural Science Foundation of China (No.31601989 and No.31672477).

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

299_2016_2068_MOESM1_ESM.jpg (896 kb)
Figure S1. Analysis of the ZjGRP promoter sequence. (JPEG 895 kb)


  1. Ambrosone A, Costa A, Leone A, Grillo S (2012) Beyond transcription: RNA-binding proteins as emerging regulators of plant response to environmental constraints. Plant Sci 182:12–18CrossRefPubMedGoogle Scholar
  2. Apel K, Hirt H (2004) Reactive oxygen species: metabolism, oxidative stress, and signal transduction. Annu Rev Plant Biol 55:373–399CrossRefPubMedGoogle Scholar
  3. Apse MP, Aharon GS, Snedden WA, Blumwald E (1999) Salt tolerance conferred by overexpression of a vacuolar Na+/H+ antiport in Arabidopsis. Science 285:1256–1258CrossRefPubMedGoogle Scholar
  4. Bandurska H (2000) Does proline accumulated in leaves of water deficit stressed barley plants confine cell membrane injury? I. Free proline accumulation and membrane injury index in drought and osmotically stressed plants. Acta Physiologiae Plantarum 22:409–415CrossRefGoogle Scholar
  5. Cervera M (2005) Histochemical and fluorometric assays for uidA (GUS) gene detection. Methods Mol Biol 286:203–214PubMedGoogle Scholar
  6. Chang Z, Liu Y, Dong H, Teng K, Han L, Zhang X (2016) Effects of cytokinin and nitrogen on drought tolerance of creeping bentgrass. PLoS One 11:e0154005CrossRefPubMedPubMedCentralGoogle Scholar
  7. Chen Y, Zong J, Tan Z, Li L, Hu B, Chen C, Chen J, Liu J (2015) Systematic mining of salt-tolerant genes in halophyte-Zoysia matrella through cDNA expression library screening. Plant Physiol Bioch 89:44–52CrossRefGoogle Scholar
  8. Demiral T, Türkan I (2006) Exogenous glycinebetaine affects growth and proline accumulation and retards senescence in two rice cultivars under NaCl stress. Environ Exp Bot 56:72–79CrossRefGoogle Scholar
  9. Du Y, Hei Q, Liu Y, Zhang H, Xu K, Xia T (2010) Isolation and characterization of a putative vacuolar Na+/H+ antiporter gene from Zoysia japonica L. J Plant Biol 53:251–258CrossRefGoogle Scholar
  10. Epstein E, Norlyn JD, Rush DW, Kingsbury RW, Kelley DB, Cunningham GA, Wrona AF (1980) Saline culture of crops: a genetic approach. Science 210:399–404CrossRefPubMedGoogle Scholar
  11. Fukutoku Y, Yamada Y (1981) Sources of proline-nitrogen in water-stressed soybean (Glycine max L.) I. Protein metabolism and proline accumulation. Plant Cell Physiol 22:1397–1404Google Scholar
  12. Ghoulam C, Foursy A, Fares K (2002) Effects of salt stress on growth, inorganic ions and proline accumulation in relation to osmotic adjustment in five sugar beet cultivars. Environ Exp Bot 47:39–50CrossRefGoogle Scholar
  13. Hidenori T, Hideki H, Shunichi K, Shinobu N, Akiko O, Akiko W, Masatsugu H, Takahiro G, Genki I, Melody M (2016) Sequencing and comparative analyses of the genomes of zoysiagrasses. DNA Res 23:171–180CrossRefGoogle Scholar
  14. Hoagland DR, Arnon DI (1950) The water-culture method for growing plants without soil. Circular California Agricultural Experiment Station, vol 347, 34 ppGoogle Scholar
  15. Huang B, Huang B, Bonos SA (2006) Breeding and genomic approaches to improving abiotic stress tolerance in plants. Plant-Environment Interactions, 3rd edn. CRC Press, Boca Raton, pp 357–376CrossRefGoogle Scholar
  16. Huang GT, Ma SL, Bai LP, Zhang L, Ma H, Jia P, Liu J, Zhong M, Guo ZF (2012) Signal transduction during cold, salt, and drought stresses in plants. Mol Biol Rep 39:969–987CrossRefPubMedGoogle Scholar
  17. Huang BR, DaCosta M, Jiang YW (2014) Research advances in mechanisms of turfgrass tolerance to abiotic stresses: from physiology to molecular biology. Crit Rev Plant Sci 33:141–189CrossRefGoogle Scholar
  18. Jia N, Liu X, Gao H (2016) A DNA2 homolog is required for DNA damage repair, cell cycle regulation, and meristem maintenance in plants. Plant Physiol 171:318–333CrossRefPubMedPubMedCentralGoogle Scholar
  19. Kim JS, Jung HJ, Lee HJ, Kim K, Goh CH, Woo Y, Oh SH, Han YS, Kang H (2008) Glycine-rich RNA-binding protein7 affects abiotic stress responses by regulating stomata opening and closing in Arabidopsis thaliana. Plant J 55:455–466CrossRefPubMedGoogle Scholar
  20. Kjelgren R, Rupp L, Kilgren D (2000) Water conservation in urban landscapes. HortScience 35:1037–1040Google Scholar
  21. Kwak KJ, Kim H-S, Jang HY, Kang H, Ahn S-J (2016) Diverse roles of glycine-rich RNA-binding protein 7 in the response of camelina (Camelina sativa) to abiotic stress. Acta Physiologiae Plantarum 38:1–11CrossRefGoogle Scholar
  22. Li C, Han L-B, Zhang X (2012) Enhanced drought tolerance of tobacco overexpressing OJERF gene is associated with alteration in proline and antioxidant metabolism. J Am Soc Hortic Sci 137:107–113CrossRefGoogle Scholar
  23. Liu C, Li S, Wang M, Xia G (2012) A transcriptomic analysis reveals the nature of salinity tolerance of a wheat introgression line. Plant Mol Biol 78:159–169CrossRefPubMedGoogle Scholar
  24. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2 − ΔΔCT method. Methods 25:402–408CrossRefPubMedGoogle Scholar
  25. Long R, Yang Q, Kang J, Zhang T, Wang H, Li M, Zhang Z (2013) Overexpression of a novel salt stress-induced glycine-rich protein gene from alfalfa causes salt and ABA sensitivity in Arabidopsis. Plant Cell Rep 32:1289–1298CrossRefPubMedGoogle Scholar
  26. Long R, Wang H, Shen Y, Kang J, Zhang T, Sun Y, Zhang Y, Li M, Yang Q (2014) Molecular cloning and functional analysis of a salt-induced gene encoding an RNA-binding protein in alfalfa. Mol Breeding 34:1465–1473CrossRefGoogle Scholar
  27. Lorkovic ZJ (2009) Role of plant RNA-binding proteins in development, stress response and genome organization. Trends Plant Sci 14:229–236CrossRefPubMedGoogle Scholar
  28. Lutts S, Kinet J, Bouharmont J (1996) Effects of salt stress on growth, mineral nutrition and proline accumulation in relation to osmotic adjustment in rice (Oryza sativa L.) cultivars differing in salinity resistance. Plant Growth Regul 19:207–218CrossRefGoogle Scholar
  29. Patton AJ, Reicher ZJ (2007) Zoysiagrass species and genotypes differ in their winter injury and freeze tolerance. Crop Sci 47:1619–1627CrossRefGoogle Scholar
  30. Petersen TN, Brunak S, von Heijne G, Nielsen H (2011) SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat Methods 8:785–786CrossRefPubMedGoogle Scholar
  31. Puyang X, An M, Han L, Zhang X (2015) Protective effect of spermidine on salt stress induced oxidative damage in two Kentucky bluegrass (Poa pratensis L.) cultivars. Ecotoxicol Environ Saf 117:96–106CrossRefPubMedGoogle Scholar
  32. Seki M, Kamei A, Yamaguchi-Shinozaki K, Shinozaki K (2003) Molecular responses to drought, salinity and frost: common and different paths for plant protection. Curr Opin Biotechnol 14:194–199CrossRefPubMedGoogle Scholar
  33. Shao HB, Liang ZS, Shao MA (2005) LEA proteins in higher plants: structure, function, gene expression and regulation. Colloids Surf, B 45:131–135CrossRefGoogle Scholar
  34. Sparkes IA, Runions J, Kearns A, Hawes C (2006) Rapid, transient expression of fluorescent fusion proteins in tobacco plants and generation of stably transformed plants. Nat Protoc 1:2019–2025CrossRefPubMedGoogle Scholar
  35. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739CrossRefPubMedPubMedCentralGoogle Scholar
  36. Tan Y, Qin Y, Li Y, Li M, Ma F (2014) Overexpression of MpGR-RBP1, a glycine-rich RNA-binding protein gene from Malus prunifolia (Willd.) Borkh., confers salt stress tolerance and protects against oxidative stress in Arabidopsis. Plant Cell. Tissue Organ C (PCTOC) 119:635–646CrossRefGoogle Scholar
  37. Teng K, Chang ZH, Xiao GZ, Guo WE, Xu LX, Chao YH, Han LB (2016a) Molecular cloning and characterization of a chlorophyll degradation regulatory gene (ZjSGR) from Zoysia japonica. Genet Mol Res. doi: 10.4238/gmr.15028176 Google Scholar
  38. Teng K, Xiao GZ, Guo WE, Yuan JB, Li J, Chao YH, Han LB (2016b) Expression of an alfalfa (Medicago sativa L.) peroxidase gene in transgenic Arabidopsis thaliana enhances resistance to NaCl and H2O2. Genet Mol Res. doi: 10.4238/gmr.15028002 Google Scholar
  39. Urano K, Kurihara Y, Seki M, Shinozaki K (2010) ‘Omics’ analyses of regulatory networks in plant abiotic stress responses. Curr Opin Plant Biol 13:132–138CrossRefPubMedGoogle Scholar
  40. Vaidyanathan H, Sivakumar P, Chakrabarty R, Thomas G (2003) Scavenging of reactive oxygen species in NaCl-stressed rice (Oryza sativa L.)—differential response in salt-tolerant and sensitive varieties. Plant Sci 165:1411–1418CrossRefGoogle Scholar
  41. Wei S, Du Z, Gao F, Ke X, Li J, Liu J, Zhou Y (2015) Global transcriptome profiles of ‘Meyer’ Zoysiagrass in response to cold stress. PLoS One 10:e0131153CrossRefPubMedPubMedCentralGoogle Scholar
  42. Yang DH, Kwak KJ, Kim MK, Park SJ, Yang KY, Kang H (2014) Expression of Arabidopsis glycine-rich RNA-binding protein AtGRP2 or AtGRP7 improves grain yield of rice (Oryza sativa) under drought stress conditions. Plant Sci 214:106–112CrossRefPubMedGoogle Scholar
  43. Zhu JK (2002) Salt and drought stress signal transduction in plants. Annu Rev Plant Biol 53:247–273CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  1. 1.Turfgrass Research Institute, College of ForestryBeijing Forestry UniversityBeijingChina
  2. 2.The College of Horticulture and GardenYangtze UniversityJingzhouChina

Personalised recommendations