Lepidium graminifolium L.: Glucosinolate Profile and Antiproliferative Potential of Volatile Isolates

Glucosinolates (GSLs) from Lepidium graminifolium L. were analyzed qualitatively and quantitatively by their desulfo-counterparts using UHPLC-DAD-MS/MS technique and by their volatile breakdown products-isothiocyanates (ITCs) using GC-MS analysis. Thirteen GSLs were identified with arylaliphatic as the major ones in the following order: 3-hydroxybenzyl GSL (glucolepigramin, 7), benzyl GSL (glucotropaeolin, 9), 3,4,5-trimethoxybenzyl GSL (11), 3-methoxybenzyl GSL (glucolimnanthin, 12), 4-hydroxy-3,5-dimethoxybenzyl GSL (3,5-dimethoxysinalbin, 8), 4-hydroxybenzyl GSL (glucosinalbin, 6), 3,4-dimethoxybenzyl GSL (10) and 2-phenylethyl GSL (gluconasturtiin, 13). GSL breakdown products obtained by hydrodistillation (HD) and CH2Cl2 extraction after hydrolysis by myrosinase for 24 h (EXT) as well as benzyl ITC were tested for their cytotoxic activity using MTT assay. Generally, EXT showed noticeable antiproliferative activity against human bladder cancer cell line UM-UC-3 and human glioblastoma cell line LN229, and can be considered as moderately active, while IC50 of benzyl ITC was 12.3 μg/mL, which can be considered as highly active.

Glucosinolate profile and Myrosinase gene expression are modulated upon Plasmodiophora brassicae infection in cabbage

Clubroot is a devastating disease of Brassicaceae caused by the biotrophic protist Plasmodiophora brassicae. The progression of clubroot disease is modulated by the glucosinolate (GSL) profile of the host plant. GSL is hydrolysed by the enzyme myrosinase upon cell disruption and gives rise to metabolites like isothiocyanate, nitriles, thiocyanates, epithionitriles and oxazolidines. Some of these metabolites play important roles in the plant’s defence mechanism. We identified 13 Myrosinase (Myro) and 28 Myrosinase-Binding Protein-like (MBP) genes from Brassica oleracea L. using a comparative genomics approach and characterised them through in silico analyses. We compared the expression patterns of these genes in a clubroot-susceptible line and a resistant line following inoculation with P. brassicae. Two BolMyro and 12 BolMBP genes were highly expressed in the susceptible line, whereas only one BolMyro and five BolMBP genes were highly expressed in the resistant line. Principal component analysis confirmed that specific GSL profiles and gene expression were modulated due to pathogen infection. Plants with higher levels of neoglucobrassicin, glucobrassicin and methooxyglucobrassicin produced disease symptoms and formed galls, whereas, plants with higher levels of sinigrin, hydroxyglucobrassicin and progoitrin produced less symptoms with almost no galls. Our results provide insights into the roles of Myro and MBP genes in GSL hydrolysis during P. brassicae infection, which will help for developing clubroot resistant cabbage lines.

Bunias erucago L.: Glucosinolate Profile and In Vitro Biological Potential

Bunias erucago belongs to the Brassicaceae family, which represents a forgotten crop of the Euro-Mediterranean area. The aim of the present study was to determine the glucosinolate profile in different plant parts and biological properties (antioxidant, anticholinesterase, and cytotoxic activities) of the isolates containing glucosinolate breakdown products. The chemical profiles were determined by using HPLC-PDA-MS/MS of desulfoglucosinolates and GC-MS of glucosinolate degradation products. The analysis of B. erucago showed the presence of seven glucosinolates: gluconapin (1), glucoraphasatin (2), glucoraphenin (3), glucoerucin (4), glucoraphanin (5), glucotropaeolin (6), and glucosinalbin (7). The total glucosinolate content ranged from 7.0 to 14.6 µmol/g of dry weight, with the major glucosinolate glucosinalbin in all parts. The antioxidant activity of all volatile isolates was not notable. At a tested concentration of 227 μg/mL, flower hydro-distillate (FH) showed good AChE inhibition, i.e., 40.9%, while root hydro-distillate (RH) had good activity against BChE, i.e., 54.3%. FH showed the best activity against both tested human bladder cancer cell lines, i.e., against T24 after 72 h, which have IC50 of 16.0 μg/mL, and against TCCSUP after 48 h with IC50 of 7.8 μg/mL, and can be considered as highly active. On the other hand, RH showed weak activity against tested cancer cells.

Coselected genes determine adaptive variation in herbivore resistance throughout the native range of Arabidopsis thaliana

The “mustard oil bomb” is a major defense mechanism in the Brassicaceae, which includes crops such as canola and the model plant Arabidopsis thaliana. These plants produce and store blends of amino acid-derived secondary metabolites called glucosinolates. Upon tissue rupture by natural enemies, the myrosinase enzyme hydrolyses glucosinolates, releasing defense molecules. Brassicaceae display extensive variation in the mixture of glucosinolates that they produce. To investigate the genetics underlying natural variation in glucosinolate profiles, we conducted a large genome-wide association study of 22 methionine-derived glucosinolates using A. thaliana accessions from across Europe. We found that 36% of among accession variation in overall glucosinolate profile was explained by genetic differentiation at only three known loci from the glucosinolate pathway. Glucosinolate-related SNPs were up to 490-fold enriched in the extreme tail of the genome-wide FST scan, indicating strong selection on loci controlling this pathway. Glucosinolate profiles displayed a striking longitudinal gradient with alkenyl and hydroxyalkenyl glucosinolates enriched in the West. We detected a significant contribution of glucosinolate loci toward general herbivore resistance and lifetime fitness in common garden experiments conducted in France, where accessions are enriched in hydroxyalkenyls. In addition to demonstrating the adaptive value of glucosinolate profile variation, we also detected long-distance linkage disequilibrium at two underlying loci, GS-OH and GS-ELONG. Locally cooccurring alleles at these loci display epistatic effects on herbivore resistance and fitness in ecologically realistic conditions. Together, our results suggest that natural selection has favored a locally adaptive configuration of physically unlinked loci in Western Europe.