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p-Coumarate:CoA ligase as a key gene in the yield of catechins in tea [Camellia sinensis (L.) O. Kuntze]

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Abstract

Tea is an important crop known for its beverage and antioxidant polyphenols—catechins and its derivatives. Catechins are synthesized through flavonoid (FL) pathway and stored in the vacuole. A metabolic flux for the operation of FL pathway is maintained through the supply of 4-coumaroyl-CoA of phenylpropanoid pathway. 4-Coumaroyl-CoA is synthesized through the catalytic activity of p-coumarate:CoA ligase (4CL) using 4-coumaric acid and acetyl-CoA as the substrates. The present manuscript reports the full-length cDNA cloning of 4CL from tea (Cs4CL accession number DQ194356) and its association with catechin yield. Cs4CL comprised of 2,165 bp with an open reading frame (ORF) of 1,764 nt, starting from 118 to 1,882 encoding 588 amino acids. Altering catechin content through a variety of environmental conditions such as drought stress (DS), abscisic acid (ABA) and gibberellic acid (GA3) treatments, and wounding established a strong positive correlation coefficient between catechins content and the expression of Cs4Cl. In addition, tea clones with high levels of catechins had higher expression of Cs4Cl whereas tea clones with lower catechins exhibited lower expression of this gene. Exposure of tea shoots to 50–100 μM catechins led to down-regulation of the expression of Cs4CL suggesting product-mediated feedback regulation and an important role for the phenylpropanoid pathway in determining catechin yield in tea.

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Fig. 1

Abbreviations

ABA:

abscisic acid

4CL:

4-coumarate CoA ligase

DS:

drought stress

FL:

flavonoid

GA3 :

gibberellic acid

PP:

phenylpropanoid

RACE:

rapid amplification of cDNA ends

RT:

reverse transcriptase

PCR:

polymerase chain reaction

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Acknowledgements

Authors are thankful to the Council of Scientific and Industrial Research (CSIR), India, for funding the present research work under the New Millennium Indian Technology Leadership Initiative (NMITLI) program entitled “Using functional genomics in plants: development and use of technologies for gene discovery and expression modulation—niche pathway engineering in tea”. KS and AR are grateful to CSIR for awarding Junior/Senior Research Fellowships. PS thanks CSIR for award of project assistantship. Technical help provided by Mr. Digvijay Singh in gene sequencing is duly acknowledged.

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Author notes

  1. Kashmir Singh

    Present address: Department of Biotechnology, Panjab University, Chandigarh, India

Authors and Affiliations

  1. Biotechnology Division, Institute of Himalayan, Bioresource Technology, P.O. Box No. 6, Palampur, 176061, India

    Arti Rani, Kashmir Singh, Payal Sood, Sanjay Kumar & P. S. Ahuja

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  1. Arti Rani
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  2. Kashmir Singh
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  3. Payal Sood
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Corresponding author

Correspondence to Sanjay Kumar.

Electronic supplementary material

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Fig. S1

Pathway for biosynthesis of flavonoids representing mainly the phenylpropanoid pathway. PAL, phenylalanine ammonia-lyase; C4H, cinnamate 4-hydroxylase; 4CL, 4-coumarate CoA ligase (PDF 15.8 KB).

Fig. S2

Sequence alignment of the putative Cs4CL and other 4CLs. The completely identical amino acids present in all sequences are marked in red, identical amino acid in most of the aligned sequences are marked in blue, and non-identical amino acids are marked in black (PDF 31.1 KB).

Fig. S3

(A) Predicted secondary structure of deduced 4-Coumarate Co A ligase (Cs4CL) by SOPMA and (B) comparison with that of At4CL. Helices, sheets, turns, and coils are indicated, respectively, with the longest, the second longest, the second shortest, and the shortest vertical lines (PDF 9.60 KB).

Fig. S4

Catechin content in leaves at different node positions (A); in response to drought stress (DS), ABA, and GA3 treatment in AB and first leaf combined (B); and in response to wounding (C) (PDF 21.7 KB).

Fig. S5

Effect of catechin on expression of Cs4CL at different time periods as shown in the figure panel (PDF 28.0 KB).

Fig. S6

Catechin content and expression of Cs4CL in tea clones with varying catechin content as analyzed by reverse northern analysis. Catechin content is shown as % g−1 dry weight in parentheses against each clone (PDF 36.6 KB).

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Rani, A., Singh, K., Sood, P. et al. p-Coumarate:CoA ligase as a key gene in the yield of catechins in tea [Camellia sinensis (L.) O. Kuntze]. Funct Integr Genomics 9, 271–275 (2009). https://doi.org/10.1007/s10142-008-0098-3

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