Plant Cell Reports

, Volume 31, Issue 9, pp 1687–1699 | Cite as

Expression of phytochelatin synthase from aquatic macrophyte Ceratophyllum demersum L. enhances cadmium and arsenic accumulation in tobacco

  • Devesh Shukla
  • Ravi Kesari
  • Seema Mishra
  • Sanjay Dwivedi
  • Rudra Deo Tripathi
  • Pravendra Nath
  • Prabodh Kumar Trivedi
Original Paper

Abstract

Phytochelatin synthase (PCS), the key enzyme involved in heavy metal detoxification and accumulation has been used from various sources to develop transgenic plants for the purpose of phytoremediation. However, some of the earlier studies provided contradictory results. Most of the PCS genes were isolated from plants that are not potential metal accumulators. In this study, we have isolated PCS gene from Ceratophyllumdemersum cv. L. (CdPCS1), a submerged rootless aquatic macrophyte, which is considered as potential accumulator of heavy metals. The CdPCS1 cDNA of 1,757 bp encodes a polypeptide of 501 amino acid residues and differs from other known PCS with respect to the presence of a number of cysteine residues known for their interaction with heavy metals. Complementation of cad1-3 mutant of Arabidopsis deficient in PC (phytochelatin) biosynthesis by CdPCS1 suggests its role in the synthesis of PCs. Transgenic tobacco plants expressing CdPCS1 showed several-fold increased PC content and precursor non-protein thiols with enhanced accumulation of cadmium (Cd) and arsenic (As) without significant decrease in plant growth. We conclude that CdPCS1 encodes functional PCS and may be part of metal detoxification mechanism of the heavy metal accumulating plant C.demersum.

Key message

Heterologous expression of PCS gene from C.demersum complements Arabidopsiscad1-3 mutant and leads to enhanced accumulation of Cd and As in transgenic tobacco.

Keywords

Arsenic Aquatic macrophyte Cadmium Ceratophyllum demersum Phytochelatin synthase Transgenic tobacco 

Abbreviations

As(V)

Arsenate

Cd

Cadmium

CdPCS1/cad1-3

cad1-3 mutant plants transformed with CdPCS1 construct

γ-EC

Gamma-glutamylcysteine

FW

Fresh weight

GSH

Glutathione

PC

Phytochelatin

PCS

Phytochelatin synthase

WT

Wild type

Supplementary material

299_2012_1283_MOESM1_ESM.doc (38 kb)
Supplementary material 1 (DOC 37 kb)
299_2012_1283_MOESM2_ESM.pptx (150 kb)
Fig. S1 Schematic representation of sequence alignment of PCS protein sequences from selected organisms such as Arabidopsis thaliana (At) and Triticumaestivum (Ta) chosen for plants; Schizosaccharomyces pombe (Sp) chosen for fungi and C. demersum (Cd) chosen as the case of present study. Black vertical bars denote cysteine residues, black vertical bars with asterisks denote conserved cysteine residues present in N-terminal. Red vertical bar with asterisks denotes conserved His-162 and blue vertical bar denotes conserved Asp-180, considered as an essential part of catalytic domain of the enzyme. Supplementary material 2 (PPT 227 kb)
299_2012_1283_MOESM3_ESM.pptx (124 kb)
Fig. S2 Fluorescence HPLC chromatograms of the mBBr-labeled plant extracts from WT and transgenic lines. Root extracts of WT and transgenic lines grown in absence of heavy metals (0 μM) were derivatized with mBBr and separated by HPLC. Peaks corresponding to cysteine, γ-EC, GSH, PC2, PC3 and PC4 standards are indicated in the chromatogram. Other unknown non-protein thiols (Peaks A-E) which are differentially accumulated in transgenic lines with respect to WT are marked. Supplementary material 3 (PPT 187 kb)
299_2012_1283_MOESM4_ESM.pptx (282 kb)
Fig. S3 Fluorescence HPLC chromatograms of the mBBr-labeled plant extracts from WT and transgenic lines. Root extracts of WT and transgenic lines exposed on Cd (200 μM) for 3 days were derivatized with mBBr and separated by HPLC. Peaks corresponding to cysteine, γ-EC, GSH, PC2, PC3 and PC4 standards are indicated in the chromatogram. Other unknown non-protein thiols (Peaks A-E) which are differentially accumulated in transgenic lines with respect to WT are marked. Supplementary material 4 (PPT 328 kb)
299_2012_1283_MOESM5_ESM.pptx (124 kb)
Fig. S4 Fluorescence HPLC chromatograms of the mBBr-labeled plant extracts from WT and transgenic lines. Root extracts of WT and transgenic lines exposed on AsV (200 μM) for 3 days were derivatized with mBBr and separated by HPLC. Peaks corresponding to cysteine, γ-EC, GSH, PC2, PC3 and PC4 standards are indicated in the chromatogram. Other unknown non-protein thiols (Peaks A-E) which are differentially accumulated in transgenic lines with respect to WT are marked. Supplementary material 5 (PPT 180 kb)

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Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Devesh Shukla
    • 1
  • Ravi Kesari
    • 1
  • Seema Mishra
    • 1
  • Sanjay Dwivedi
    • 1
  • Rudra Deo Tripathi
    • 1
  • Pravendra Nath
    • 1
  • Prabodh Kumar Trivedi
    • 1
  1. 1.CSIR-National Botanical Research InstituteLucknowIndia

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