Proteome alterations monitored by DIGE analysis in Silybum marianum cell cultures elicited with methyl jasmonate and methyl B cyclodextrin.

UNLABELLED: Elicitation with methyl jasmonate (MeJA) or/and cyclodextrin (CD) strongly induced silymarin (Sm) accumulation in suspensions of Silybum marianum, with most of Sm isomers being detected in the culture medium. This induction provides a model platform to characterize the regulation of flavonolignan accumulation and release in response to elicitors and, with this aim, changes in the S. marianum cell proteome were investigated. The DIGE technique was used to detect statistically significant changes in the cell's proteome. A total number of 1269 unique spots were detected, 67 of which were de-regulated upon elicitation. Nineteen spots were identified by nLC-MS/MS database search analysis. Identified proteins belong to a few categories, including metabolism, stress and defense responses and transport processes. The most abundant group was represented by pathogenesis-related (PR) proteins and heat shock proteins. Two proteins related to transport process were identified and both were upregulated by elicitation. One was identified as Ras-related protein Rab11C of the Rab family of small ATPase superfamily. A second protein was identified as an ABC transporter. Some of the identified proteins are discussed with respect to their putative role in the extracellular flavonolignan accumulation in S. marianum cultures.

BIOLOGICAL SIGNIFICANCE: Most approaches to increase secondary metabolite yields using plant cell cultures have been focused on the optimization of its biosynthesis. The study of other post biosynthetic events, like chemical or enzymatic modifications, transport, storage/secretion and catabolism/degradation are also biotechnologically relevant. Secretion is of particular interest since if cell cultures are to be used routinely for the commercial production, they must release the targeted metabolites into the extracellular medium. Elicitor-induced silymarin accumulation and release in S. marianum cell cultures provide a responsive model system to profile both alterations in proteins related to monolignol/flavonoid biosynthesis and to identify potential systems involved in secretion of secondary metabolites. The proteomic approach undertaken in this work has permitted identify some of the events occurring in elicited S. marianum cell cultures. One attainment of this study is that a vesicular transport mechanism could be involved in the release of this class of secondary metabolites to the extracellular compartment. This finding forms a baseline for future research on a non-sequenced medicinal plant S. marianum at molecular level.