ATML1 Regulates the Differentiation of ER Body-containing Large Pavement Cells in Rosette Leaves of Brassicaceae Plants.

Endoplasmic reticulum (ER)-derived organelles, ER bodies, participate in the defense against herbivores in Brassicaceae plants. ER bodies accumulate ß-glucosidases, which hydrolyse specialized thioglucosides known as glucosinolates to generate bioactive substances. In Arabidopsis thaliana, the leaf ER (LER) bodies are formed in large pavement cells, which are found in the petioles, margins, and blades of rosette leaves. However, the regulatory mechanisms involved in establishing large pavement cells are unknown. Here, we show that the ARABIDOPSIS THALIANA MERISTEM L1 LAYER (ATML1) transcription factor regulates the formation of LER bodies in large pavement cells of rosette leaves. Overexpression of ATML1 enhanced the expression of LER body-related genes and the number of LER body-containing large pavement cells, whereas its knockout resulted in opposite effects. ATML1 enhances endoreduplication and cell size through LOSS OF GIANT CELLS FROM ORGANS (LGO). Although the overexpression and knockout of LGO affected the appearance of large pavement cells in Arabidopsis, the effect on LER body-related gene expression and LER body formation was weak. LER body-containing large pavement cells were also found in Eutrema salsugineum, another Brassicaceae species. Our results demonstrate that ATML1 establishes large pavement cells to induce LER body formation in Brassicaceae plants, and thereby possibly contributes to the defense against herbivores.

Loss of MYB34 Transcription Factor Supports the Backward Evolution of Indole Glucosinolate Biosynthesis in a Subclade of the Camelineae Tribe and Releases the Feedback Loop in This Pathway in Arabidopsis.

Glucosinolates are specialized defensive metabolites characteristic of the Brassicales order. Among them, aliphatic and indolic glucosinolates (IGs) are usually highly abundant in species from the Brassicaceae family. The exceptions this trend are species representing a subclade of the Camelineae tribe, including Capsella and Camelina genera, which have reduced capacity to produce and metabolize IGs. Our study addresses the contribution of specific glucosinolate-related myeloblastosis (MYB) transcription factors to this unprecedented backward evolution of IG biosynthesis. To this end, we performed phylogenomic and functional studies of respective MYB proteins. The obtained results revealed weakened conservation of glucosinolate-related MYB transcription factors, including loss of functional MYB34 protein, in the investigated species. We showed that the introduction of functional MYB34 from Arabidopsis thaliana partially restores IG biosynthesis in Capsella rubella, indicating that the loss of this transcription factor contributes to the backward evolution of this metabolic pathway. Finally, we performed an analysis of the impact of particular myb mutations on the feedback loop in IG biosynthesis, which drives auxin overproduction, metabolic dysregulation and strong growth retardation caused by mutations in IG biosynthetic genes. This uncovered the unique function of MYB34 among IG-related MYBs in this feedback regulation and consequently in IG conservation in Brassicaceae plants.

Evolutionary changes in the glucosinolate biosynthetic capacity in species representing Capsella, Camelina and Neslia genera.

Glucosinolates are unique thioglucosides that evolved in the order Brassicales. These compounds function in plant adaptation to the environment, including combating plant pathogens, herbivore deterrence and abiotic stress tolerance. In line with their defensive functions glucosinolates usually accumulate constitutively in relatively high amounts in all tissues of Brassicaceae plants. Here we performed glucosinolate analysis in different organs of selected species representing Capsella, Camelina and Neslia genera, which similarly as the model plant Arabidopsis thaliana belong to the Camelineae tribe. We also identified orthologs of A. thaliana glucosinolate biosynthetic genes in the published genomes of some of the investigated species. Subsequent gene expression and phylogenetic analyses enabled us an insight into the evolutionary changes in the transcription of these genes and in the sequences of respective proteins that occurred within the Camelineae tribe. Our results indicated that glucosinolates are highly abundant in siliques and roots of the investigated species but hardly, if at all, produced in leaves. In addition to this unusual tissular distribution we revealed reduced structural diversity of methionine-derived aliphatic glucosinolates (AGs) with elevated accumulation of rare long chain AGs. This preference seems to correlate with evolutionary changes in genes encoding methylthioalkylmalate synthases that are responsible for the elongation of AG side chains. Finally, our results indicate that the biosynthetic pathway for tryptophan-derived indolic glucosinolates likely lost its main functions in immunity and resistance towards sucking insects and is on its evolutionary route to be shut off in the investigated species.

Glutathione S-Transferases in the Biosynthesis of Sulfur-Containing Secondary Metabolites in Brassicaceae Plants.

Plants in the Brassicaceae family have evolved the capacity to produce numerous unique and structurally diverse sulfur-containing secondary metabolites, including constitutively present thio-glucosides, also known as glucosinolates, and indole-type phytoalexins, which are induced upon pathogen recognition. Studies on the glucosinolate and phytoalexin biosynthetic pathways in the model plant Arabidopsis thaliana have shown that glutathione donates the sulfur atoms that are present in these compounds, and this further suggests that specialized glutathione S-transferases (GSTs) are involved in the biosynthesis of glucosinolates and sulfur-containing phytoalexins. In addition, experimental evidence has shown that GSTs also participate in glucosinolate catabolism. Several candidate GSTs have been suggested based on co-expression analysis, however, the function of only a few of these enzymes have been validated by enzymatic assays or with phenotypes of respective mutant plants. Thus, it remains to be determined whether biosynthesis of sulfur-containing metabolites in Brassicaceae plants requires specific or nonspecific GSTs.