Skip to main content

Ectopic expression of PgRab7 in rice plants (Oryza sativa L.) results in differential tolerance at the vegetative and seed setting stage during salinity and drought stress

Protoplasma volume 254, pages 109–124 (2017)Cite this article

Abstract

In this work, we have overexpressed a vesicle trafficking protein, Rab7, from a stress-tolerant plant, Pennisetum glaucum, in a high-yielding but stress-sensitive rice variety Pusa Basmati-1 (PB-1). The transgenic rice plants were tested for tolerance against salinity and drought stress. The transgenic plants showed considerable tolerance at the vegetative stage against both salinity (200 mM NaCl) and drought stress (up to 12 days after withdrawing water). The protection against salt and drought stress may be by regulating Na+ ion homeostasis, as the transgenic plants showed altered expression of multiple transporter genes, including OsNHX1, OsNHX2, OsSOS1, OsVHA, and OsGLRs. In addition, decreased generation and maintenance of lesser reactive oxygen species (ROS), with maintenance of chloroplast grana and photosynthetic machinery was observed. When evaluated for reproductive growth, 89–96 % of seed setting was maintained in transgenic plants during drought stress; however, under salt stress, a 33–53 % decrease in seed setting was observed. These results indicate that PgRab7 overexpression in rice confers differential tolerance at the seed setting stage during salinity and drought stress and could be a favored target for raising drought-tolerant crops.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

43,34 €

Price includes VAT (Finland)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10

Abbreviations

DAB:

3,3-diaminobenzidine

ETR:

electron transport rate

IRGA:

infrared gas analyzer

MS:

Murashige and Skoog

MV:

methyl viologen

qRT-PCR:

quantitative real time-PCR

ROS:

reactive oxygen species

TEM:

transmission electron microscopy

UV-light:

ultra violet light

WT:

wild-type

References

  • Agarwal PK, Agarwal P, Jain P, Jha B, Reddy MK, Sopory SK (2008) Constitutive overexpression of a stress-inducible small GTP binding protein PgRab7 from Pennisetum glaucum enhances abiotic stress tolerance in transgenic tobacco. Plant Cell Rep 27:105–115

    CAS  Article  PubMed  Google Scholar 

  • Apse MP, Aharon GS, Snedden WA, Blumwald E (1999) Salt tolerance conferred by overexpression of a vacuolar Na+/H+ antiport in Arabidopsis. Science 258:1256–1258

    Article  Google Scholar 

  • Arnon DI (1949) Copper enzymes in isolated chloroplasts. Poly-phenoloxidase in beta vulgaris. Plant Physiol 24:1–15

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Baisakh N, RamanaRao MV, Rajasekaran K, Subudhi P, Janda J, Galbraith D, Vanier C, Pereira A (2012) Enhanced salt stress tolerance of rice plants expressing a vacuolar H+-ATPase subunit c1 (SaVHAc1) gene from the halophyte grass Spartina alterniflora Loisel. Plant Biotechnol J 10:453–464

    CAS  Article  PubMed  Google Scholar 

  • Balderhaar HJK, Arlt H, Ostrowicz C, Bröcker C, Sündermann F, Brandt R, Babst M, Ungermann C (2010) The Rab GTPase Ypt7 is linked to retromer-mediated receptor recycling and fusion at the yeast late endosome. J Cell Sci 123:4085–4094

    CAS  Article  PubMed  Google Scholar 

  • Bassil E, Ohto MA, Esumi T, Tajima H, Zhu Z, Cagnac O, Belmonte M, Peleg Z, Yamaguchi T, Blumwald E (2011) The Arabidopsis intracellular Na+/H+ antiporters NHX5 and NHX6 are endosome associated and necessary for plant growth and development. Plant Cell 23:224–239

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Bassil E, Coku A, Blumwald E (2012) Cellular ion homeostasis: emerging roles of intracellular NHX Na+/H+ antiporters in plant growth and development. J Exp Bot 63:5727–5740

    CAS  Article  PubMed  Google Scholar 

  • Bolte S, Schiene K, Dietz KJ (2000) Characterization of a small GTP binding protein of the rab 5 family in Mesembryanthemum crystallinum with increased level of expression during early salt stress. Plant Mol Biol 42:923–936

    CAS  Article  PubMed  Google Scholar 

  • Botha CEJ, Aoki N, Scofield GN, Liu L, Furbank RT, White RG (2008) A xylem sap retrieval pathway in rice leaf blades: evidence of a role for endocytosis? J Exp Bot 59:2945–2954

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    CAS  Article  PubMed  Google Scholar 

  • Bruckert F, Laurent O, Satre M (2000) Rab7, a multifaceted GTP-binding protein regulating access to degradative compartments in eukaryotic cells. Protoplasma 210:108–116

    CAS  Article  Google Scholar 

  • Bucci C, Thomsen P, Nicoziani P, McCarthy J, van Deurs B (2000) Rab7: a key to lysosome biogenesis. Mol Biol Cell 11:467–480

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Cheong MS, Yun DJ (2007) Salt-stress signaling. J Plant Biol 50:148–155

    CAS  Article  Google Scholar 

  • Cheung AY, de Vries SC (2008) Membrane trafficking: intracellular highways and country roads. Plant Physiol 147:1451–1453

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Cruz de Carvalho MH (2008) Drought stress and reactive oxygen species: production, scavenging and signaling. Plant Signal Behav 3:156–165

    Article  PubMed  PubMed Central  Google Scholar 

  • Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation: version II. Plant Mol Biol Rep 1:19–21

    CAS  Article  Google Scholar 

  • Deunff EL, Davoine C, Dantec CL, Billard JP, Huault C (2004) Oxidative burst and expression of germin/oxo genes during wounding of ryegrass leaf blades: comparison with senescence of leaf sheaths. Plant J 38:421–431

    Article  PubMed  Google Scholar 

  • Diaz-Vivancos P, Faize M, Barba-Espin G, Faize L, Petri C, Hernández JA, Burgos L (2013) Ectopic expression of cytosolic superoxide dismutase and ascorbate peroxidase leads to salt stress tolerance in transgenic plums. Plant Biotechnol J 11:976–985

    CAS  Article  PubMed  Google Scholar 

  • Duan XG, Yang AF, Gao F, Zhang SL, Zhang JR (2007) Heterologous expression of vacuolar H(+)-PPase enhances the electrochemical gradient across the vacuolar membrane and improves tobacco cell salt tolerance. Protoplasma 232:87–95

    CAS  Article  PubMed  Google Scholar 

  • Fedoroff N (2006) Redox regulatory mechanisms in cellular stress responses. Ann Bot 98:289–300

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Garg AK, Kim JK, Owens TG, Ranwala AP, Choi YD, Kochian LV, Wu RJ (2002) Trehalose accumulation in rice plants confers high tolerance levels to different abiotic stresses. Proc Natl Acad Sci U S A 99:15898–15903

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • George S, Parida A (2011) Over-expression of a Rab family GTPase from phreatophyte Prosopis juliflora confers tolerance to salt stress on transgenic tobacco. Mol Biol Rep 38:1669–1674

    CAS  Article  PubMed  Google Scholar 

  • Gill SS, Tuteja N (2010) Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants. Plant Physiol Biochem 48:909–930

    CAS  Article  PubMed  Google Scholar 

  • Hamaji K, Nagira M, Yoshida K, Ohnishi M, Oda Y, Uemura T, Goh T, Sato MH, Morita MT, Tasaka M, Hasezawa SI, Nakano A, Nishimura IH, Maeshima M, Fukaki H, Mimura T (2009) Dynamic aspects of ion accumulation by vesicle traffic under salt stress in Arabidopsis. Plant Cell Physiol 50:2023–2033

    CAS  Article  PubMed  Google Scholar 

  • Hasegawa PM, Bressan RA, Zhu JK, Bohnert HJ (2000) Plant cellular and molecular responses to high salinity. Annu Rev Plant Physiol Plant Mol Biol 51:463–499

    CAS  Article  PubMed  Google Scholar 

  • Jia Y, Yang X, Feng Y, Jilani G (2008) Differential response of root morphology to potassium deficient stress among rice genotypes varying in potassium efficiency. J Zhejiang Univ Sci B 9:427–434

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Kader MA, Seidel T, Golldack D, Lindberg S (2006) Expressions of OsHKT1, OsHKT2, and OsVHA are differentially regulated under NaCl stress in salt-sensitive and salt-tolerant rice (Oryza sativa L.) cultivars. J Exp Bot 57:4257–4268

  • Kawano N, Ito O, Sakagami J (2009) Morphological and physiological responses of rice seedlings to complete submergence (flash flooding). Ann Bot 103:161–169

    Article  PubMed  Google Scholar 

  • Kieffer F, Simon-Plas F, Maume BF, Blein JP (1997) Tobacco cells contain a protein, immunologically related to the neutrophil small G protein Rac2 and involved in elicitor induced oxidative burst. FEBS Lett 403:149–153

    CAS  Article  PubMed  Google Scholar 

  • Levine A, Belenghi B, Damari-Weisler H, Granot D (2001) Vesicle associated membrane protein of Arabidopsis suppresses bax-induced apoptosis in yeast downstream of oxidative burst. J Biol Chem 276:46284–46289

    CAS  Article  PubMed  Google Scholar 

  • Leyman B, Geelen D, Blatt MR (1999) Localization and control of expression of NtSyr1, a tobacco SNARE protein. Plant J 24:369–381

    Article  Google Scholar 

  • Limpens E, Ivanov S, van Esse W, Voets G, Fedorova E, Bisseling T (2009) Medicago N2-fixing symbiosomes acquire the endocytic identity marker Rab7 but delay the acquisition of vacuolar identity. Plant Cell 21:2811–2828

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Liu F, Guo J, Bai P, Duan Y, Wang X, Cheng Y, Feng H, Huang L, Kang Z (2012) Wheat TaRab7 GTPase is part of the signaling pathway in responses to stripe rust and abiotic stimuli. PLoS ONE 7(5):e37146

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-delta delta C(T)) method. Methods 25:402–408

    CAS  Article  PubMed  Google Scholar 

  • Mazel A, Leshem Y, Tiwari BS, Levine A (2004) Induction of salt and osmotic stress tolerance by overexpression of an intracellular vesicle trafficking protein AtRab7 (AtRabG3e). Plant Physiol 134:118–128

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Mimura T, Kura-Hotta M, Tsujimura T, Ohnishi M, Miura M, Okazaki Y, Mimura M, Maeshima M, Washitani-Nemoto S (2003) Rapid increase of vacuolar volume in response to salt stress. Planta 216:397–402

    CAS  PubMed  Google Scholar 

  • Munns R (2002) Comparitive physiology of salt and water stress. Plant Cell Environ 25:239–250

    CAS  Article  PubMed  Google Scholar 

  • Munns R, Tester M (2008) Mechanisms of salinity tolerance. Ann Rev Plant Biol 59:651–681

    CAS  Article  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassay with tissue culture. Physiol Plant 15:473–476

    CAS  Article  Google Scholar 

  • Nahm MY, Kim SW, Yun D, Lee SY, Cho MJ, Bahk JD (2003) Molecular and biochemical analyses of OsRab7, a rice Rab7 homolog. Plant Cell Physiol 44:1341–1349

    CAS  Article  PubMed  Google Scholar 

  • Niu X, Bressan RA, Hasegawa PM, Pardo JM (1995) Ion homeostasis in NaCl stress environments. Plant Physiol 109:735–742

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • O’Mahony PJ, Oliver MJ (1999) Characterization of a desiccation-responsive small GTP-binding protein (Rab2) from the desiccation-tolerant grass Sporobolus stapfianus. Plant Mol Biol 39:809–821

    Article  PubMed  Google Scholar 

  • Park M, Lee H, Lee JS, Byun MO, Kim BG (2009) In planta measurements of Na+ using fluorescent dye corona green. J Plant Biol 52:298–302

    CAS  Article  Google Scholar 

  • Park JY, Kim YY, Martinoia E, Lee Y (2008) Long-distance transporters of inorganic nutrients in plants. J Plant Biol 51:240–247

    CAS  Article  Google Scholar 

  • Radomiljac JD, Whelan J, van der Merwe M (2013) Coordinating metabolite changes with our perception of plant abiotic stress responses: emerging views revealed by integrative-omic analyses. Metabolites 3:761–786

    Article  PubMed  PubMed Central  Google Scholar 

  • Randall SK, Crowell DN (1999) Protein isoprenylation in plants. Crit Rev Biochem Mol Biol 34:325–338

    CAS  Article  PubMed  Google Scholar 

  • Rosales KR, Peralta ER, Wong SY, Edinger AL (2009) Rab7 activation by growth factor withdrawal contributes to the induction of apoptosis. Mol Biol Cell 20:2831–2840

    CAS  Article  Google Scholar 

  • Schimmoller F, Reizman H (1993) Involvement of Ypt7p, a small GTPase, in traffic from late endosome to the vacuole in yeast. J Cell Sci 106:823–830

    PubMed  Google Scholar 

  • Shi H, Lee BH, Wu SJ, Zhu JK (2003) Overexpression of a plasma membrane Na+/H+ antiporter gene improves salt tolerance in Arabidopsis thaliana. Nat Biotechnol 21:81–85

    CAS  Article  PubMed  Google Scholar 

  • Singh S, Ambastha V, Levine A, Sopory SK, Yadava PK, Tripathy BC, Tiwari BS (2015) Anhydrobiosis and programmed cell death in plants: commonalities and differences. Curr Plant Biol 2:12–20

    Article  Google Scholar 

  • Singla-Pareek SL, Reddy MK, Sopory SK (2003) Genetic engineering of the glyoxalase pathway in tobacco leads to enhanced salinity tolerance. Proc Natl Acad Sci U S A 100:14672–14677

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Singla-Pareek SL, Pareek A, Sopory SK (2007) Transgenic plants for dry and saline environments. In: Jenks MA et al (ed) Advances in molecular breeding toward drought and salt tolerant crops. Springer, pp 501-530

  • Stenmark H (2009) Rab GTPases as coordinators of vesicle traffic. Nat Rev 10:513–525

    CAS  Article  Google Scholar 

  • Sui JM, Li R, Fan QC, Song L, Zheng CH, Wang JS, Qiao LX, Yu SL (2013) Isolation and characterization of a stress responsive small GTP-binding protein AhRabG3b in peanut (Arachis hypogaea L.). Euphytica 189:161–172

    CAS  Article  Google Scholar 

  • Tripathy MK, Tyagi W, Goswami M, Kaul T, Singla-Pareek SL, Deswal R, Reddy MK, Sopory SK (2012) Characterization and functional validation of tobacco PLC delta for abiotic stress tolerance. Plant Mol Biol Rep 30:488–497

  • Verma D, Singla-Pareek SL, Rajagopal D, Reddy MK, Sopory SK (2007) Functional validation of a novel isoform of Na+/H+ antiporter from Pennisetum glaucum for enhancing salinity tolerance in rice. J Biosci 32:621–628

    CAS  Article  PubMed  Google Scholar 

  • Wu SJ, Ding L, Zhu JK (1996) SOS1, a genetic locus essential for salt tolerance and potassium acquisition. Plant Cell 8:617–627

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Yoshida S, Forno DA, Cock JH, Gomez KA (1972) Laboratory manual for physiological studies of rice. IRRI, Manila, Philippines

    Google Scholar 

  • Zerial M, McBride H (2001) Rab proteins as membrane organizers. Nat Rev Mol Cell Biol 2:107–117

    CAS  Article  PubMed  Google Scholar 

  • Zhang HX, Blumwald E (2001) Transgenic salt tolerant tomato plants accumulate salt in the foliage but not in the fruits. Nat Biotechnol 19:765–768

    CAS  Article  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by a grant from the Department of Biotechnology, New Delhi, Government of India to SKS. We are thankful to Mr. S. C. P. Sharma and Dr. G. Saini (Jawaharlal Nehru University, New Delhi, India) for their help during TEM study. We thank Prof. S. J. Roux (The University of Texas at Austin, Texas, USA) for his help in correcting the manuscript.

Author information

Author notes

  1. Manas Kumar Tripathy

    Present address: Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX, 78712, USA

  2. Sudhir K. Sopory

    Present address: Jawaharlal Nehru University, New Delhi, 110067, India

Affiliations

  1. Plant Molecular Biology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, 110067, India

    Manas Kumar Tripathy, Malireddy K. Reddy & Sudhir K. Sopory

  2. School of Life Sciences, Jawaharlal Nehru University, New Delhi, 110067, India

    Budhi Sagar Tiwari

  3. Department of Botany, University of Delhi, Delhi, 110007, India

    Renu Deswal

Authors
  1. Manas Kumar Tripathy
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Budhi Sagar Tiwari
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Malireddy K. Reddy
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Renu Deswal
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Sudhir K. Sopory
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Manas Kumar Tripathy or Sudhir K. Sopory.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Handling Editor: Bhumi Nath Tripathi

Electronic supplementary material

Below is the link to the electronic supplementary material.

figure 11

High resolution image.

Table S1

List of genes studied by real time PCR assay.

Fig. S1

Representative pictures to show relative salinity tolerance and morphology of wild-type and PgRab7 -transgenic plants. Transgenic plants and WT seedlings were grown under control (A and E), 100 mM NaCl (B and F), 175 mM NaCl (C and G), and 200 mM NaCl (D and H). Pictures showing (A, B, C, D) the number of seed germination (15-day-old seedlings), (E, F, G, H) shoot and root length (10-day-old seedlings), respectively, of WT and transgenic seedlings subjected to salt stress as mentioned above.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Tripathy, M.K., Tiwari, B.S., Reddy, M.K. et al. Ectopic expression of PgRab7 in rice plants (Oryza sativa L.) results in differential tolerance at the vegetative and seed setting stage during salinity and drought stress. Protoplasma 254, 109–124 (2017). https://doi.org/10.1007/s00709-015-0914-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00709-015-0914-2

Keywords

  • PgRab7
  • Rice
  • Salinity
  • Drought
  • Membrane trafficking