Endoplasmic reticulum stress controls PIN-LIKES abundance and thereby growth adaptation.

Extreme environmental conditions eventually limit plant growth [J. R. Dinneny, Annu. Rev. Cell Dev. Biol. 35, 1-19 (2019), N. Gigli-Bisceglia, C. Testerink, Curr. Opin. Plant Biol. 64, 102120 (2021)]. Here, we reveal a mechanism that enables multiple external cues to get integrated into auxin-dependent growth programs in Arabidopsis thaliana. Our forward genetics approach on dark-grown hypocotyls uncovered that an imbalance in membrane lipids enhances the protein abundance of PIN-LIKES (PILS) [E. Barbez et al., Nature 485, 119 (2012)] auxin transport facilitators at the endoplasmic reticulum (ER), which thereby limits nuclear auxin signaling and growth rates. We show that this subcellular response relates to ER stress signaling, which directly impacts PILS protein turnover in a tissue-dependent manner. This mechanism allows PILS proteins to integrate environmental input with phytohormone auxin signaling, contributing to stress-induced growth adaptation in plants.

Should I stay or should I go: the functional importance and regulation of lipid diffusion in biological membranes.

Biological membranes are highly dynamic, in particular due to the constant exchange of vesicles between the different compartments of the cell. In addition, the dynamic nature of membranes is also caused by their inherently fluid properties, with the diffusion of both proteins and lipids within their leaflets. Lipid diffusion is particularly difficult to study in vivo but recent advances in optical microscopy and lipid visualization now enable the characterization of lipid lateral motion, and here we review these methods in plants. We then discuss the parameters that affect lipid diffusion in membranes and explore their consequences on the formation of membrane domains at different scales. Finally, we consider how controlled lipid diffusion affects membrane functions during cell signaling, development, and environmental interactions.

Author Correction: Light triggers PILS-dependent reduction in nuclear auxin signalling for growth transition.

A Correction to this paper has been published: https://doi.org/10.1038/s41477-021-00924-y.

FRUITFULL Is a Repressor of Apical Hook Opening in Arabidopsis thaliana.

Plants adjust their architecture to a constantly changing environment, requiring adaptation of differential growth. Despite their importance, molecular switches, which define growth transitions, are largely unknown. Apical hook development in dark grown Arabidopsis thaliana (A. thaliana) seedlings serves as a suitable model for differential growth transition in plants. Here, we show that the phytohormone auxin counteracts the light-induced growth transition during apical hook opening. We, subsequently, identified genes which are inversely regulated by light and auxin. We used in silico analysis of the regulatory elements in this set of genes and subsequently used natural variation in gene expression to uncover correlations between underlying transcription factors and the in silico predicted target genes. This approach uncovered that MADS box transcription factor AGAMOUS-LIKE 8 (AGL8)/FRUITFULL (FUL) modulates apical hook opening. Our data shows that transient FUL expression represses the expression of growth stimulating genes during early phases of apical hook development and therewith guards the transition to growth promotion for apical hook opening. Here, we propose a role for FUL in setting tissue identity, thereby regulating differential growth during apical hook development.

The Road to Auxin-Dependent Growth Repression and Promotion in Apical Hooks.

The phytohormone auxin controls growth rates within plant tissues, but the underlying mechanisms are still largely enigmatic. The apical hook is a superb model to understand differential growth, because it displays both auxin-dependent growth repression and promotion. In this special issue on membranes, we illustrate how the distinct utilization of vesicle trafficking contributes to the spatial control of polar auxin transport, thereby pinpointing the site of growth repression in apical hooks. We moreover highlight that the transition to growth promotion is achieved by balancing inter- and intracellular auxin transport. We emphasize here that the apical hook development is a suitable model to further advance our mechanistic knowledge on plant growth regulation.

Light triggers PILS-dependent reduction in nuclear auxin signalling for growth transition.

The phytohormone auxin induces or represses growth depending on its concentration and the underlying tissue type. However, it remains unknown how auxin signalling is modulated to allow tissues transiting between repression and promotion of growth. Here, we used apical hook development as a model for growth transitions in plants. A PIN-FORMED (PIN)-dependent intercellular auxin transport module defines an auxin maximum that is causal for growth repression during the formation of the apical hook. Our data illustrate that growth transition for apical hook opening is largely independent of this PIN module, but requires the PIN-LIKES (PILS) putative auxin carriers at the endoplasmic reticulum. PILS proteins reduce nuclear auxin signalling in the apical hook, leading to the de-repression of growth and the onset of hook opening. We also show that the phytochrome (phy) B-reliant light-signalling pathway directly regulates PILS gene activity, thereby enabling light perception to repress nuclear auxin signalling and to control growth. We propose a novel mechanism, in which PILS proteins allow external signals to alter tissue sensitivity to auxin, defining differential growth rates.

Histochemical Staining of ß-Glucuronidase and Its Spatial Quantification.

Microscope images of plant specimens showing expression of GUS markers, besides being very beautiful, provide useful information regarding various biological processes. However, the information extracted from these images is often purely qualitative, and in many publications is not subjected to quantification. Here, we describe a very simple quantification method for GUS histochemical staining that enables detection of subtle differences in gene expression at cellular, tissue, or organ level. The quantification method described is based on the freely available image analysis software ImageJ that is widely used by the scientific community. We exemplify the method by quantifying small and precise changes (at the cellular level) as well as broad changes (at the organ level) in the expression of two previously published reporter lines, such as the pPILS2::GUS and pPILS5::GUS. The method presented here represents an easy tool for converting visual information from GUS histochemical staining images into quantifiable data and is of general importance for plant biologists performing GUS activity-based evaluation of reporter genes.

A novel putative auxin carrier family regulates intracellular auxin homeostasis in plants.

The phytohormone auxin acts as a prominent signal, providing, by its local accumulation or depletion in selected cells, a spatial and temporal reference for changes in the developmental program. The distribution of auxin depends on both auxin metabolism (biosynthesis, conjugation and degradation) and cellular auxin transport. We identified in silico a novel putative auxin transport facilitator family, called PIN-LIKES (PILS). Here we illustrate that PILS proteins are required for auxin-dependent regulation of plant growth by determining the cellular sensitivity to auxin. PILS proteins regulate intracellular auxin accumulation at the endoplasmic reticulum and thus auxin availability for nuclear auxin signalling. PILS activity affects the level of endogenous auxin indole-3-acetic acid (IAA), presumably via intracellular accumulation and metabolism. Our findings reveal that the transport machinery to compartmentalize auxin within the cell is of an unexpected molecular complexity and demonstrate this compartmentalization to be functionally important for a number of developmental processes.

Dihydrosphingosine-induced programmed cell death in tobacco BY-2 cells is independent of H2O2 production.

Sphinganine or dihydrosphingosine (d18:0, DHS), one of the most abundant free sphingoid Long Chain Base (LCB) in plants, has been recently shown to induce both cytosolic and nuclear calcium transient increases and a correlated Programmed Cell Death (PCD) in tobacco BY-2 cells. In this study, in order to get deeper insight into the LCB signaling pathway leading to cell death, the putative role of Reactive Oxygen Species (ROS) has been investigated. We show that DHS triggers a rapid dose-dependent production of H2O2 that is blocked by diphenyleniodonium (DPI), indicating the involvement of NADPH oxidase(s) in the process. In addition, while DPI does not block DHS-induced calcium increases, the ROS production is inhibited by the broad spectrum calcium channel blocker lanthanum (La³+). Therefore, ROS production occurs downstream of DHS-induced Ca²+ transients. Interestingly, DHS activates expression of defense-related genes that is inhibited by both La³+ and DPI. Since DPI does not prevent DHS-induced cell death, these results strongly indicate that DHS-induced H2O2 production is not implicated in PCD mechanisms but rather would be associated to basal cell defense mechanisms.