Induction of resveratrol biosynthesis in Vitis amurensis cells by heterologous expression of the Arabidopsis constitutively active, Ca2+-independent form of the AtCPK1 gene

• Published in: Process biochemistry (1991) 2017-03, Vol.54, p.144-155
• Main Authors: Veremeichik, G.N., Grigorchuk, V.P., Shkryl, Y.N., Bulgakov, D.V., Silantieva, S.A., Bulgakov, V.P.
• Format: Article
• Language: English
• Subjects: 4-Coumarate-CoA ligases; Bioengineering; Biosynthesis; Calcium; Calcium-dependent protein kinase; Callus; Callus culture; Cell activation; Cell growth; Chromatography; Gene expression; High performance liquid chromatography; Liquid chromatography; Metabolic engineering; Metabolites; Protein interaction; Resveratrol; Signal transduction; Spectrometry; Transduction; Vitis amurensis
• ISSN: 1359-5113; 1873-3298
• DOI: 10.1016/j.procbio.2016.12.026

[Display omitted] •Mutant isoforms of the AtCPK1 gene were overexpressed in V. amurensis calli.•Constitutively active isoform of the AtCPK1 gene stimulates resveratrol biosynthesis.•Effect of AtCPK1 is an indirect process and is a part of general defense reactions.•The work may be helpful in metabolic engineering of plant secondary products. In the present study, we established transgenic calli of Vitis amurensis that expressed a constitutively active, Ca2+-independent form of the AtCPK1 gene (AtCPK1-Ca) and calli with a non-active form of the gene. High-performance liquid chromatography with UV and high-resolution mass-spectrometry revealed that the predominant metabolites synthesized in our transgenic callus cultures were trans-resveratrol di-glucoside, trans-piceid, trans-resveratrol, trans-ε-viniferin and trans-δ-viniferin. The resveratrol content in the AtCPK1-Ca-transformed callus cultures exceeded that in the control cultures up to 90-fold. Furthermore, the expression of the AtCPK1-Ca gene caused cell growth activation, which led to the enhancement of resveratrol production up to 137 times that of the control calli (69.7mgL−1 vs. 0.51mgL−1). Real-time PCR analysis showed that AtCPK1-Ca overexpression caused increasing of the expression of the key enzymes of phenylpropanoid pathway of resveratrol biosynthesis, 4-coumarate-CoA ligases. Thus, heterologous expression of constitutively active CDPK genes can be used to bioengineer plant cell cultures that produce stilbenes. Possible mechanisms for AtCPK1-mediated signal transduction were proposed by the reconstruction of known protein–protein interactions within CPK1-assotiated protein modules.