Induction of anthraquinone biosynthesis in Rubia cordifolia cells by heterologous expression of a calcium‐dependent protein kinase gene

Abstract

Calcium-dependent protein kinases (CDPKs) play an important role in plant cell responses to stress and pathogenic attack. In this study, we investigated the effect of heterologous expression of the Arabidopsis CDPK gene, AtCPK1, on anthraquinone production in transgenic Rubia cordifolia cells. AtCPK1 variants (a constitutively active, Ca 2þ -independent form and a non-active form used as a negative control) were transferred to callus cells by agrobacterial transformation. Overexpression of the constitutively active, Ca 2þ -independent form in R. cordifolia cells caused a 10-fold increase in anthraquinone content compared with non-transformed control cells, while the nonactive form of AtCPK1 had no effect on anthraquinone production. Real-time PCR measurements showed that the activation of anthraquinone biosynthesis in transgenic calli correlated with the activation of isochorismate synthase gene expression. The activator effect of AtCPK1 was stable during prolonged periods of transgenic cell cultivation (more than 3 years) and the transgenic cultures exhibited high growth. Our results provide the first evidence that a CDPK gene can be used for the engineering of secondary metabolism in plant cells.

Application of Agrobacterium Rol Genes in Plant Biotechnology: A Natural Phenomenon of Secondary Metabolism Regulation

“…In a new research, an Arabidopsis CDPK gene was expressed in R. cordifolia cells. Constitutive expression of the gene caused significant activation of anthraquinone biosynthesis (Shkryl et al, 2011), thus confirming our expectation that CDPKs are directly involved in secondary metabolism regulation.…”

Induced effect of Na+ on ganoderic acid biosynthesis in static liquid culture of Ganoderma lucidum via calcineurin signal transduction

“…Further work such as knocking down cam , cna , crz1 , ena1 , and Ca 2+ ‐ATPase genes in G. lucidum with RNAi or using a mass spectrometry‐based approach to identify the signal‐inducible proteins (Chen et al, 2005) would help clarify the more detailed role of each calcineurin signal target in GA biosynthesis regulation. In addition, as reported in Rubia cordifolia plant cells for production of anthraquinone (Shkryl et al, 2011), heterologous expression of calcium sensors may improve cellular secondary metabolism, which could be an interesting future topic for the GA applied research.…”

TherolBGene Suppresses Reactive Oxygen Species in Transformed Plant Cells through the Sustained Activation of Antioxidant Defense

“…This effect is combined with the necrotizing effect of rolB. Because these features are associated with a high ROS level Shkryl et al, 2011), one would expect that the gene could induce ROS production. Our results, however, contradict this hypothesis.…”