Plant Sterol Clustering Correlates with Membrane Microdomains as Revealed by Optical and Computational Microscopy

Local inhomogeneities in lipid composition play a crucial role in the regulation of signal transduction and membrane traffic. This is particularly the case for plant plasma membrane, which is enriched in specific lipids, such as free and conjugated forms of phytosterols and typical phytosphingolipids. Nevertheless, most evidence for microdomains in cells remains indirect, and the nature of membrane inhomogeneities has been difficult to characterize. We used a new push–pull pyrene probe and fluorescence lifetime imaging microscopy (FLIM) combined with all-atom multiscale molecular dynamics simulations to provide a detailed view on the interaction between phospholipids and phytosterol and the effect of modulating cellular phytosterols on membrane-associated microdomains and phase separation formation. Our understanding of the organization principles of biomembranes is limited mainly by the challenge to measure distributions and interactions of lipids and proteins within the complex environment of living cells. Comparing phospholipids/phytosterol compositions typical of liquid-disordered (Ld) and liquid-ordered (Lo) domains, we furthermore show that phytosterols play crucial roles in membrane homeostasis. The simulation work highlights how state-of-the-art modeling alleviates some of the prior concerns and how unrefuted discoveries can be made through a computational microscope. Altogether, our results support the role of phytosterols in the lateral structuring of the PM of plant cells and suggest that they are key compounds for the formation of plant PM microdomains and the lipid-ordered phase.

Phytocytokines function as immunological modulators of plant immunity

Plant plasma membrane-resident immune receptors regulate plant immunity by recognizing microbe-associated molecular patterns (MAMPs), damage-associated molecular patterns (DAMPs), and phytocytokines. Phytocytokines are plant endogenous peptides, which are usually produced in the cytosol and released into the apoplast when plant encounters pathogen infections. Phytocytokines regulate plant immunity through activating an overlapping signaling pathway with MAMPs/DAMPs with some unique features. Here, we highlight the current understanding of phytocytokine production, perception and functions in plant immunity, and discuss how plants and pathogens manipulate phytocytokine signaling for their own benefits during the plant-pathogen warfare.

Nep1-like proteins as a target for plant pathogen control

The lack of efficient methods to control the major diseases of crops most important to agriculture leads to huge economic losses and seriously threatens global food security. Many of the most important microbial plant pathogens, including bacteria, fungi, and oomycetes, secrete necrosis- and ethylene-inducing peptide 1 (Nep1)-like proteins (NLPs), which critically contribute to the virulence and spread of the disease. NLPs are cytotoxic to eudicot plants, as they disturb the plant plasma membrane by binding to specific plant membrane sphingolipid receptors. Their pivotal role in plant infection and broad taxonomic distribution makes NLPs a promising target for the development of novel phytopharmaceutical compounds. To identify compounds that bind to NLPs from the oomycetes Pythium aphanidermatum and Phytophthora parasitica, a library of 587 small molecules, most of which are commercially unavailable, was screened by surface plasmon resonance. Importantly, compounds that exhibited the highest affinity to NLPs were also found to inhibit NLP-mediated necrosis in tobacco leaves and Phytophthora infestans growth on potato leaves. Saturation transfer difference-nuclear magnetic resonance and molecular modelling of the most promising compound, anthranilic acid derivative, confirmed stable binding to the NLP protein, which resulted in decreased necrotic activity and reduced ion leakage from tobacco leaves. We, therefore, confirmed that NLPs are an appealing target for the development of novel phytopharmaceutical agents and strategies, which aim to directly interfere with the function of these major microbial virulence factors. The compounds identified in this study represent lead structures for further optimization and antimicrobial product development.

Exogenous Fatty Acid Hydroperoxide Perception as Elicitor Is Related to Modulation of Plant Plasma Membrane Structure

Fatty acid hydroperoxides (HPOs) are amphiphilic molecules naturally produced by plants in stressed conditions and involved in plant immunity as signalling molecules. Although some studies report their potential use as exogenous biocontrol agents for plant protection, evaluation of their efficiency in planta is lacking and no information is available about their mechanism of action. In this work, the potential of two HPO forms, 13-HPOD and 13-HPOT, as plant defence elicitors and the underlying mechanism of action are investigated. Both HPOs trigger Arabidopsis innate immunity. They increase plant resistance to the pathogenic fungi Botrytis cinerea and activate early immunity-related defence responses, like ROS production. As our previous study has suggested that HPOs are able to interact with the plant plasma membrane (PPM) lipid fraction, we have further investigated the effects of HPOs on biomimetic PPM structure using complementary biophysics tools. Results show that HPO insertion into PPM impacts its global structure without solubilizing it. 13-HPOT, with an additional double bond compared to 13-HPOD, exerts a higher effect by fluidifying and reducing the thickness of the bilayer. Correlation between biological assays and biophysical analysis suggests that lipid amphiphilic elicitors that directly act on membrane lipids might trigger early plant defence events.

A plant plasma-membrane H+-ATPase promotes yeast TORC1 activation via its carboxy-terminal tail

The Target of Rapamycin Complex 1 (TORC1) involved in coordination of cell growth and metabolism is highly conserved among eukaryotes. Yet the signals and mechanisms controlling its activity differ among taxa, according to their biological specificities. A common feature of fungal and plant cells, distinguishing them from animal cells, is that their plasma membrane contains a highly abundant H+-ATPase which establishes an electrochemical H+ gradient driving active nutrient transport. We have previously reported that in yeast, nutrient-uptake-coupled H+ influx elicits transient TORC1 activation and that the plasma-membrane H+-ATPase Pma1 plays an important role in this activation, involving more than just establishment of the H+ gradient. We show here that the PMA2 H+-ATPase from the plant Nicotiana plumbaginifolia can substitute for Pma1 in yeast, to promote H+-elicited TORC1 activation. This H+-ATPase is highly similar to Pma1 but has a longer carboxy-terminal tail binding 14–3–3 proteins. We report that a C-terminally truncated PMA2, which remains fully active, fails to promote H+-elicited TORC1 activation. Activation is also impaired when binding of PMA2 to 14–3–3 s is hindered. Our results show that at least some plant plasma-membrane H+-ATPases share with yeast Pma1 the ability to promote TORC1 activation in yeast upon H+-coupled nutrient uptake.