Conversion of Exogenous Cholesterol into Glycoalkaloids in Potato Shoots, Using Two Methods for Sterol Solubilisation

Abstract

Steroidal glycoalkaloids (SGA) are toxic secondary metabolites naturally occurring in the potato, as well as in certain other Solanaceous plant species, such as tomato, eggplant and pepper. To investigate the steroidal origin of SGA biosynthesis, cut potato shoots were fed cholesterol labelled with deuterium (D) in the sterol ring structure (D5- or D6-labelled), or side chain (D7-labelled), and analysed after three or five weeks. The labelled cholesterol and presence of D-labelled SGA were analysed by GC-MS and LC-MS/MS, respectively. When feeding D-labelled cholesterol solubilised in Tween-80, labelled cholesterol in free form became present in both leaves and stems, although the major part was recovered as steryl esters. Minor amounts of D-labelled SGA (α-solanine and α-chaconine) were identified in cholesterol-treated shoots, but not in blank controls, or in shoots fed D6-27-hydroxycholesterol. Solubilising the labelled cholesterol in methyl-β-cyclodextrin instead of Tween-80 increased the levels of labelled SGA up to 100-fold, and about 1 mole% of the labelled cholesterol was recovered as labelled SGA in potato leaves. Both side chain and ring structure D labels were retained in SGA, showing that the entire cholesterol molecule is converted to SGA. However, feeding side chain D7-labelled cholesterol resulted in D5-labelled SGA, indicating that two hydrogen atoms were released during formation of the SGA nitrogen-containing ring system. Feeding with D7-sitosterol did not produce any labelled SGA, indicating that cholesterol is a specific SGA precursor. In conclusion, we have demonstrated a superior performance of methyl-β-cyclodextrin for delivery of cholesterol in plant tissue feeding experiments, and given firm evidence for cholesterol as a specific sterol precursor of SGA in potato.

Weighted Gene Co-expression Network Analysis of the Dioscin Rich Medicinal Plant Dioscorea nipponica

“…The biosynthetic pathways of cholestenol saponins, such as the important phytohormones sitosterol and brassinosteroids ( Symons and Reid, 2004 ), have been intensively studied ( Chu et al, 2005 ), while the biosynthesis of diosgenin, a spimstanol saponin, remains poorly understood. Previous work using isotope labeling has suggested that spimstanol saponins are derived from cholesterol ( Bennett and Heftmann, 1965 ; Tomita and Uomori, 1971 ), and reports have increasingly shown that cholesterol is the precursor of the C24-desmethyl sterols ( McCue et al, 2005 , 2006 , 2007 ; Itkin et al, 2011 , 2013 ; Petersson et al, 2013 ), providing further evidence to support the notion that dioscin is derived from cholesterol ( Figure 2 ). C24 methylation is the main divergence point between the biosynthesis of C24-desmethyl sterols and cholestenol saponins, and this process can be regulated by MeJA ( Ren et al, 2009 ).…”

Changes in the Plant β-Sitosterol/Stigmasterol Ratio Caused by the Plant Parasitic Nematode Meloidogyne incognita

“…It seems plausible that the observed changes in the sterol pool are linked to a metabolic reaction against the infection by M. incognita . For example, in solanaceous plants, cholesterol can make up a significant portion of the overall sterol pool and has been suggested as a precursor of toxic steroidal alkaloids and glycoalkaloids [ 31 ]. Campesterol is used in numerous plants as precursor for the synthesis of brassinosteroid phytohormones, essential for the regulation of numerous plant processes, such as cell expansion and elongation, senescence and protection against drought and chilling [ 32 ].…”

Transcript profiling of two potato cultivars during glycoalkaloid-inducing treatments shows differential expression of genes in sterol and glycoalkaloid metabolism

“…S3 . Sterols in experiments with transgenic plants were quantified by GC-MS using desmosterol, cycloeucalenol and 24-methylene cycloartanol as added internal standards for different sterol fractions 13 .…”

“…SGAs are derived from the isoprenoid pathway, and the sterol cholesterol has been identified as a metabolic precursor 11 12 13 ( Supplementary Fig. S1 ).…”