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Transcript profiling of terpenoid indole alkaloid pathway genes and regulators reveals strong expression of repressors in Catharanthus roseus cell cultures

  • Cell Biology and Morphogenesis
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Abstract

The understanding of the complexities and molecular events regulating genes and the activators involved in terpenoid indole alkaloid (TIA) metabolism is known to a certain extent in cell cultures of an important TIA yielding plant, Catharanthus roseus, though it is not yet complete. Recently, the repressors of early TIA pathway genes have also been identified. However, their roles in the regulation of TIA pathway in C. roseus cell cultures remains yet unknown. We have made a comparative profiling of genes catalyzing the important steps of 2-C methyl-D-erythritol-4-phosphate (MEP), shikimate and TIA biosynthetic pathways, their activator and repressors using macroarray, semiquantitative RT-PCR and northern analyses in a rotation culture system of C. roseus comprising differentiated and proliferated cells. Our results demonstrate that TIA biosynthetic pathway genes and their activators show variable expression pattern, which was correlated with the changes in the cellular conditions in these systems. Under similar conditions, TIA pathway repressors show strong and consistent expression. The role of repressors in the complex regulation of the TIA pathway in C. roseus cell cultures is discussed. The results were supported by HPLC data, which demonstrated that the molecular program of cellular differentiation is intimately linked with TIA pathway gene expression and TIA production in C. roseus cell cultures.

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Abbreviations

BAP:

Benzylamino purine

CCC:

Compact callus cluster

2, 4-D:

2, 4-Dichlorophenoxy acetic acid

HPLC:

High performance liquid chromatography

MeJA:

Methyl ester of jasmonic acid

MEP:

2-C-methyl-D-erythritol-4-phosphate

ORCA:

Octadecanoid-derivative responsive Catharanthus AP2-domain protein

R-CCC:

Reverted-compact callus cluster

RT-PCR:

Reverse transcriptase-polymerase chain reaction

S:

Suspension

TIA:

Terpenoid indole alkaloid

YE:

Yeast extract

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Acknowledgments

AD acknowledges Council of Scientific and Industrial Research (CSIR), Government of India for the award of senior research fellowship. This work was supported by the Department of Biotechnology, Government of India.

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Authors and Affiliations

  1. National Centre for Plant Genome Research, Jawaharlal Nehru University Campus, P. O. Box 10531, Aruna Asaf Ali Marg, New Delhi, 110 067, India

    Ajaswrata Dutta, Digvijay Singh, Sushil Kumar & Jayanti Sen

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  1. Ajaswrata Dutta
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  2. Digvijay Singh
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  3. Sushil Kumar
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  4. Jayanti Sen
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Correspondence to Ajaswrata Dutta.

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Communicated by P. Lakshmanan.

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299_2007_305_MOESM1_ESM.doc

299_2007_305_Fig7_ESM.jpg

Fig. S1 Gene expression analysis with macroarray system. (a) Designed image of the cDNAmacroarray. 50ng of denatured purified PCR product spotted on Hybond N membrane and hybridized with [α-32P] labeled leaf induced (b) CCC, (c) S and (d) R-CCC mRNA probes. CrActin and NptII were used as positive and negative controls respectively. (JPG 33.4 kb)

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Dutta, A., Singh, D., Kumar, S. et al. Transcript profiling of terpenoid indole alkaloid pathway genes and regulators reveals strong expression of repressors in Catharanthus roseus cell cultures. Plant Cell Rep 26, 907–915 (2007). https://doi.org/10.1007/s00299-007-0305-z

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  • DOI: https://doi.org/10.1007/s00299-007-0305-z

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