Skip to main content
Log in

Gene expression in cadmium-tolerant Datura innoxia: detection and characterization of cDNAs induced in response to Cd2+

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

The response of a metal tolerant plant to heavy metal stress involves a number of biochemical pathways. To investigate the overall molecular response of a metal-tolerant plant to heavy-metal exposure, suppressive subtractive hybridization was used to create a library enriched in cadmium-induced cDNAs from cadmium-tolerant Datura innoxia. Two differential screening steps were used to screen the cadmium-induced library resulting in 8 putative cadmium-specific cDNAs out of a pool of 94 clones. Reverse transcriptase polymerase chain reaction was used to confirm that 4 of these 8 clones were cadmium-specific, while the other 4 were induced under heat shock or in the no treatment cells in addition to cadmium exposure. All 8 cDNAs were sequenced and used to search for identification against GenBank. One of the 4 cadmium-specific cDNAs had homology to a sulfur transferase-family protein in Arabidopsis thaliana. The possible link between this result and the heavy-metal response of plants is discussed.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Al-Lahham, A., Rohde, V. and Zimmerman, M. 1999. Biosynthesis of phytochelatins in the fission yeast. Phytochelatin synthesis: a second role for the glutathione synthetase gene of Schizosaccharomyces pombe. Yeast 15: 385–396.

    Google Scholar 

  • Altschul, S.F., Gish, W., Miller, W., Myers, E.W. and Lipman, D.J. 1990. Basic local alignment search tool. J. Mol. Biol. 215: 403–410.

    Google Scholar 

  • Chen, J., Zhou, J. and Goldsbrough, P.B. 1997. Characterization of phytochelatin synthase from tomato. Physiol. Plant. 101: 165–172.

    Google Scholar 

  • Clemens, S. 2001. Molecular mechanisms of plant metal tolerance and homeostasis. Planta 212: 475–486.

    Google Scholar 

  • Clemens, S., Kim, E.J., Neumann, D., Schroeder, J.I. 1999. Tolerance to toxic metals by a gene family of phytochelatin synthases from plants and yeast EMBO J. 18: 3325–3333.

    Google Scholar 

  • Cobbett, C. 2000. Phytochelatins and their roles in heavy metal detoxification. Plant Physiol. 123: 825–832.

    Google Scholar 

  • Cobbett, C. and Goldsbrough, P. 2002. Phytochelatins and metallothioneins: roles in heavy metal detoxification and homeostasis. Annu. Rev. Plant Biol. 53: 159–182.

    Google Scholar 

  • de Knecht, J.A., van Dillen, M., Koevoets, P.L.M., Schat, H., Verkleij, J.A.C. and Ernst, W.H.O. 1994. Phytochelatins in cadmium-sensitive and cadmium-tolerant Silene vulgaris. Plant Physiol. 104: 255–261.

    Google Scholar 

  • Delhaize, E., Robinson, N.J. and Jackson, P.J. 1989. Effects of cadmium on gene expression in cadmium-tolerant and cadmiumsensitive Datura innoxia cells. Plant Mol. Biol. 12: 487–497.

    Google Scholar 

  • Diatchenko, L., Chenchik, A. and Siebert, P.D. 1998. Suppression subtractive hybridization: a method of generating subtracted cDNA libraries starting from poly(A+) or total RNA. In: P.D. Siebert and J.W. Larrick (Eds.) RT-PCR Methods for Gene Cloning and Analysis, BioTechniques Books, MA, pp. 213–239.

  • Diatchenko, L., Lau,Y.-F.C., Campbell, A., Chenchik, A., Moqadam, F., Huang, B., Lukyanov, S., Lukyanov, K., Gurskaya, N., Sverdlow, E.D. and Siebert, P.D. 1996. Suppression subtractive hybridization: a method for generating differentially regulated or tissue-specific cDNA probes and libraries. Proc. Natl. Acad. Sci. USA 93: 6025–6030.

    Google Scholar 

  • Edelman, L., Czarnecka, E. and Key, J.L. 1986. Induction and accumulation of heat shock-specific poly(A+) RNAs and proteins in soybean seedlings during arsenite and cadmium treatments. Plant Physiol. 86: 1048–1056.

    Google Scholar 

  • Grill, E., Loffler, S., Winnacker, E.-L. and Zenk, M.H. 1989. Phytochelatins, the heavy-metal-binding peptides of plants, are synthesized from glutathione by a specific γ-glutamylcysteine dipeptidyl transpeptidase (phytochelatin synthase). Proc. Natl. Acad. Sci. USA 86: 6838–6842.

    Google Scholar 

  • Ha, S.B., Smith, A.P., Howden, R., Dietrich, W.M., Bugg, S., O'Connell, M.J., Goldsbrough, P.B. and Cobbett, C.S. 1999. Phytochelatin synthase genes from Arabidopsis and the yeast, Schizosaccharomyces pombe. Plant Cell 11: 1153–1164.

    Google Scholar 

  • Heiss, S., Schäfer, H.J., Haag-Kerwer, A. and Rausch, T. 1999. Cloning sulfur assimilation genes of Brassica juncea L.: cadmium differentially affects the expression of a putative lowaffinity sulfate transporter and isoforms of ATP sulfurylase and APS reductase. Plant Mol. Biol. 39: 847–857.

    Google Scholar 

  • Jackson, P.J., Roth, E.J., McClure, P.R. and Naranjo, C.M. 1984. Selection, isolation and characterization of cadmium-resistant Datura innoxia suspension cultures. Plant Physiol. 75: 914–918.

    Google Scholar 

  • Jackson, P.J., Delhaize, E. and Kuske, C.R. 1992. Biosynthesis and metabolic roles of cadystins (γ-EC) nG and their precursors in Datura innoxia. Plant Soil 146: 281–289.

    Google Scholar 

  • Kärenlampi, S., Schat, H., Vangronsveld, J., Verkleij, J.A.C., van der Lelie, D., Mergeay, M. and Tervahauta, A.I. 2000. Genetic engineering in the improvement of plants for phytoremediation of metal polluted soils. Environ. Poll. 107: 225–231.

    Google Scholar 

  • Leustek, T., Martin, M.N., Bick, J.-A. and Davies, J.P. 2000. Pathways and regulation of sulfur metabolism revealed through molecular and genetic studies. Annu. Rev. Plant Physiol. Plant Mol. Biol. 51: 141–165.

    Google Scholar 

  • Louie, M. 2000. Cadmium-induced gene expression in Datura innoxia. Master thesis, San Francisco State University, San Francisco, CA.

    Google Scholar 

  • Lukyanov, K.A., Matz, M.V., Bogdanova, E.A., Gurskaya, N.G. and Lukyanov, S.A. 1996. Molecule by molecule PCR amplification of complex DNA mixtures for direct sequencing: an approach to in vitro cloning. Nucl. Acids Res. 24: 2194–2195.

    Google Scholar 

  • Macnair, M.R. 1993. The genetics of metal tolerance in vascular plants. New Phytol. 124: 541–559

    Google Scholar 

  • Moniz de Sa, M. and Drounin, G. 1996. Phylogeny and substitution rates of angiosperm actin genes. Mol. Biol. Evol. 13: 1198–1212.

    Google Scholar 

  • Nakamura, T., Yamaguchi, Y. and Sano, H. 2000. Plant mercaptopyruvate sulfurtranferases. Molecular cloning, subcellular localization and enzymatic activities. Eur. J. Biochem. 267: 5621–5630.

    Google Scholar 

  • Neumann, D., Litchtenberger, O., Gunther, D., Tschiersch, K. and Nover, L. 1994. Heat-shock proteins induce heavy metal tolerance in higher plants. Planta 194: 360–367.

    Google Scholar 

  • Ow, D.W. 1996. Heavy metal tolerance genes: prospective tools for bioremediation. Resources Conserv. Recycl. 18: 135–149.

    Google Scholar 

  • Papenbrock, J. and Schmidt, A. 2000a. Characterization of a sulfurtransferase from Arabidopsis thaliana. Eur. J. Biochem. 267: 145–154.

    Google Scholar 

  • Papenbrock, J. and Schmidt, A. 2000b. Characterization of two sulfurtransferase isozymes from Arabidopsis thaliana. Eur. J. Biochem. 267: 5571–5579.

    Google Scholar 

  • Robinson, N.J., Ratliff, R.L., Anderson, P.J., Delhaize, E., Berger, J.M. and Jackson, P.J. 1988. Biosynthesis of poly(γ-glutamylcysteinyl)glycines in cadmium-tolerant Datura innoxia (Mill.) cells. Plant Sci. 56: 197–204.

    Google Scholar 

  • Sambrook, J., Fritsch, E.F. and Maniatis, T. 1989. Molecular Cloning: A Laboratory Manual, 2nd ed.. Cold Spring Harbor Laboratory Press, Plainview, NY.

    Google Scholar 

  • Schmidt, A. and Jäger, K. 1992. Open questions about sulfur metabolism in plants. Annu. Rev. Plant Physiol. Plant Mol. Biol. 43: 325–349.

    Google Scholar 

  • Tomsett, A.B. and Thurman, D.A. 1988. Molecular biology of metal tolerances of plants. Plant Cell Environ. 11: 383–394.

    Google Scholar 

  • Urwin, P.E., Groom, Q.J., Robinson, N.J. 1996. Characterization of two cDNAs and identification of two proteins that accumulate in response to cadmium in cadmium-tolerant Datura innoxia (Mill.) cells. J. Exp. Bot. 47: 1019–1024.

    Google Scholar 

  • Vatamaniuk, O.K., Mari, S., Lu, Y.P. and Rea, P.A. 1999.AtPCS1, a phytochelatin synthase from Arabidopsis: isolation and in vitro reconstitution. Proc. Natl. Acad. Sci. USA 96: 7110–7115.

    Google Scholar 

  • Wang, Z. and Brown, D.D. 1991. A gene expression screen. Proc. Natl. Acad. Sci. USA 88: 11505–11509.

    Google Scholar 

  • Yan, S.L., Tsay, C.C. and Chen, Y.R. 2000. Isolation and characterization of phytochelatin synthase in rice seedlings. Proc. Natl. Sci. Council ROC 24: 202–207.

    Google Scholar 

  • Zenk, M.H. 1996. Heavy metal detoxificaiton in higher plants - a review. Gene 179: 21–30.

    Google Scholar 

  • Zhu, Y.L., Pilon-Smits, E.A.H., Jouanin, L. and Terry, N. 1999a. Overexpression of glutathione synthetase in indian mustard enhances cadmium accumulation and tolerance. Plant Physiol. 119: 73–80.

    Google Scholar 

  • Zhu, Y.L., Pilon-Smits, E.A.H., Tarun, A.S., Weber, S.U., Jouanin, L., Terry, N. 1999b. Cadmium tolerance and accumulation in Indian mustard is enhanced by overexpressing γ-glutamylcysteine synthetase. Plant Physiol. 121: 1169–1177.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Jane G. DeWitt.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Louie, M., Kondor, N. & DeWitt, J.G. Gene expression in cadmium-tolerant Datura innoxia: detection and characterization of cDNAs induced in response to Cd2+ . Plant Mol Biol 52, 81–89 (2003). https://doi.org/10.1023/A:1023926225931

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1023926225931

Navigation