Pseudophosphorylation of Arabidopsis jasmonate biosynthesis enzyme lipoxygenase 2 via mutation of Ser600 inhibits enzyme activity.

Jasmonates are oxylipin phytohormones critical for plant resistance against necrotrophic pathogens and chewing herbivores. An early step in their biosynthesis is catalyzed by non-heme iron lipoxygenases (LOX; EC 1.13.11.12). In Arabidopsis thaliana, phosphorylation of Ser600 of AtLOX2 was previously reported, but whether phosphorylation regulates AtLOX2 activity is unclear. Here, we characterize the kinetic properties of recombinant WT AtLOX2 (AtLOX2WT). AtLOX2WT displays positive cooperativity with α-linolenic acid (α-LeA, jasmonate precursor), linoleic acid (LA), and arachidonic acid (AA) as substrates. Enzyme velocity with endogenous substrates α-LeA and LA increased with pH. For α-LeA, this increase was accompanied by a decrease in substrate affinity at alkaline pH; thus, the catalytic efficiency for α-LeA was not affected over the pH range tested. Analysis of Ser600 phosphovariants demonstrated that pseudophosphorylation inhibits enzyme activity. AtLOX2 activity was not detected in phosphomimics Atlox2S600D and Atlox2S600M when α-LeA or AA were used as substrates. In contrast, phosphonull mutant Atlox2S600A exhibited strong activity with all three substrates, α-LeA, LA, and AA. Structural comparison between the AtLOX2 AlphaFold model and a complex between 8R-LOX and a 20C polyunsaturated fatty acid suggests a close proximity between AtLOX2 Ser600 and the carboxylic acid head group of the polyunsaturated fatty acid. This analysis indicates that Ser600 is located at a critical position within the AtLOX2 structure and highlights how Ser600 phosphorylation could affect AtLOX2 catalytic activity. Overall, we propose that AtLOX2 Ser600 phosphorylation represents a key mechanism for the regulation of AtLOX2 activity and, thus, the jasmonate biosynthesis pathway and plant resistance.

Plant SUMO E3 Ligases: Function, Structural Organization, and Connection With DNA.

Protein modification by the small ubiquitin-like modifier (SUMO) plays an important role in multiple plant processes, including growth, development, and the response to abiotic stresses. Mechanistically, SUMOylation is a sequential multi-enzymatic process where SUMO E3 ligases accelerate SUMO conjugation while also influencing target identity and interactions. This review explores the biological functions of plant SUMO E3 ligases [SAP AND MIZ1 DOMAIN-CONTAINING LIGASE (SIZs), METHYL METHANESULFONATE-SENSITIVITY PROTEIN 21 (MMS21s), and PROTEIN INHIBITOR OF ACTIVATED STAT-LIKE (PIALs)] in relation to their molecular activities and domains. We also explore the sub-cellular localization of SUMO E3 ligases and review evidence suggesting a connection between certain SUMO E3 ligases and DNA that contributes to gene expression regulation.

Toxic Responses of Palladium Accumulation in Duckweed (Lemna minor): Determination of Biomarkers.

Palladium (Pd) is a trace metal of the platinum group elements, representing an emerging contaminant for the environment. It is of great interest to characterize the bioaccumulation and toxicity of Pd to improve our toxicological knowledge for this contaminant. Under standardized toxicity testing conditions, we analyzed Pd accumulation and toxicity effects on the duckweed Lemna minor exposed to nominal concentrations from 2 to 50 µM. The inhibitory effect was significant (p < 0.05) from 8 µM of Pd, starting with 9.5% of growth inhibition and a decrease of 1 cm for the root size. Under 12.5 µM of Pd, the bioaccumulated Pd of 63.93 µg/g fresh weight inhibited plant growth by 37.4%, which was caused by a strong oxidative stress in the cytosol and organelles containing DNA. Under 25 and 50 µM of Pd, bioaccumulated Pd was able to deteriorate the entire plant physiology including chlorophyll synthesis, the photosystem II antenna complex, and the photochemical reactions of photosynthesis. In fact, plants treated with 50 µM Pd accumulated Pd up to 255.95 µg/g fresh weight, causing a strong decrease in total biomass and root elongation process. Therefore, we showed several growth, physiological, and biochemical alterations which were correlated with the bioaccumulation of Pd. These alterations constituted toxicity biomarkers of Pd with different lowest-observed-effect dose, following this order: root size = growth inhibition < catalase activity = carotenoid content = reactive oxygen species production = total thiols < chlorophyll a/b = variable fluorescence to maximal fluorescence intensity ratio = absorbed-light energy transfer from the chlorophyll a antenna to the photosystem II reaction center = performance index of photosystem II activity < VJ . Therefore, the present study provides insight into the toxicity mechanism of Pd in L. minor plants under standardized testing conditions. Environ Toxicol Chem 2021;40:1630-1638. © 2021 SETAC.