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All plant cells are surrounded by a cell wall that provides cohesion, protection, and a means of directional growth to plants. Cellulose microfibrils contribute the main biomechanical scaffold for most of these walls. The biosynthesis of cellulose, which typically is the most prominent constituent of the cell wall and therefore Earth's most abundant biopolymer, is finely attuned to developmental and environmental cues. Our understanding of the machinery that catalyzes and regulates cellulose biosynthesis has substantially improved due to recent technological advances in, for example, structural biology and microscopy. Here, we provide a comprehensive overview of the structure, function, and regulation of the cellulose synthesis machinery and its regulatory interactors. We aim to highlight important knowledge gaps in the field, and outline emerging approaches that promise a means to close those gaps.

International audience; Anionic phospholipids, which include phosphatidic acid, phosphatidylserine, and phosphoinositides, represent a small percentage of membrane lipids. They are able to modulate the physical properties of membranes, such as their surface charges, curvature, or clustering of proteins. Moreover, by mediating interactions with numerous membrane-associated proteins, they are key components in the establishment of organelle identity and dynamics. Finally, anionic lipids also act as signaling molecules, as they are rapidly produced or interconverted by a set of dedicated enzymes. As such, anionic lipids are major regulators of many fundamental cellular processes, including cell signaling, cell division, membrane trafficking, cell growth, and gene expression. In this review, we describe the functions of anionic lipids from a cellular perspective. Using the localization of each anionic lipid and its related metabolic enzymes as starting points, we summarize their roles within the different compartments of the endomembrane system and address their associated developmental and physiological consequences.

Summary Plants are able to orient their growth according to gravity, which ultimately controls both shoot and root architecture.1 Gravitropism is a dynamic process whereby gravistimulation induces the asymmetric distribution of the plant hormone auxin, leading to asymmetric growth, organ bending, and subsequent reset of auxin distribution back to the original pre-gravistimulation situation.1, 2, 3 Differential auxin accumulation during the gravitropic response depends on the activity of polarly localized PIN-FORMED (PIN) auxin-efflux carriers.1, 2, 3, 4 In particular, the timing of this dynamic response is regulated by PIN2,5,6 but the underlying molecular mechanisms are poorly understood. Here, we show that MEMBRANE ASSOCIATED KINASE REGULATOR2 (MAKR2) controls the pace of the root gravitropic response. We found that MAKR2 is required for the PIN2 asymmetry during gravitropism by acting as a negative regulator of the cell-surface signaling mediated by the receptor-like kinase TRANSMEMBRANE KINASE1 (TMK1).2,7, 8, 9, 10 Furthermore, we show that the MAKR2 inhibitory effect on TMK1 signaling is antagonized by auxin itself, which triggers rapid MAKR2 membrane dissociation in a TMK1-dependent manner. Our findings suggest that the timing of the root gravitropic response is orchestrated by the reversible inhibition of the TMK1 signaling pathway at the cell surface., Graphical Abstract, Highlights • MAKR2 is co-expressed with PIN2 and regulates the pace of root gravitropism • MAKR2 controls PIN2 asymmetric accumulation at the root level during gravitropism • MAKR2 binds to and is a negative regulator of the TMK1 receptor kinase • Auxin antagonizes the MAKR2 inhibition of TMK1 by delocalizing MAKR2 in the cytosol, Marquès-Bueno, Armengot et al. show that the unstructured protein MAKR2 controls the dynamics of the root gravitropic response by acting as a negative regulator of the TMK1 receptor kinase. In addition, the MAKR2 inhibitory effect on TMK1 signaling is antagonized by auxin itself, which triggers rapid MAKR2 membrane dissociation in a TMK1-dependent manner.

Phosphoinositides are low-abundant lipids that participate in the acquisition of membrane identity through their spatiotemporal enrichment in specific compartments. Phosphatidylinositol 4-phosphate (PI4P) accumulates at the plant plasma membrane driving its high electrostatic potential, and thereby facilitating interactions with polybasic regions of proteins. PI4Kα1 has been suggested to produce PI4P at the plasma membrane, but how it is recruited to this compartment is unknown. Here, we pin-point the mechanism that tethers Arabidopsis thaliana phosphatidylinositol 4-kinase alpha1 (PI4Kα1) to the plasma membrane via a nanodomain-anchored scaffolding complex. We established that PI4Kα1 is part of a complex composed of proteins from the NO-POLLEN-GERMINATION, EFR3-OF-PLANTS, and HYCCIN-CONTAINING families. Comprehensive knockout and knockdown strategies revealed that subunits of the PI4Kα1 complex are essential for pollen, embryonic, and post-embryonic development. We further found that the PI4Kα1 complex is immobilized in plasma membrane nanodomains. Using synthetic mis-targeting strategies, we demonstrate that a combination of lipid anchoring and scaffolding localizes PI4Kα1 to the plasma membrane, which is essential for its function. Together, this work opens perspectives on the mechanisms and function of plasma membrane nanopatterning by lipid kinases., PI4Kα1 is targeted to plasma membrane nanodomains by a lipid-anchored heterotetrameric complex essential for plant cell survival, including gametophytic, embryonic, and post-embryonic development.

Phosphoinositides are low-abundant lipids that participate in the acquisition of membrane identity through their spatiotemporal enrichment in specific compartments. PI4P accumulates at the plant plasma membrane driving its high electrostatic potential, and thereby facilitating interactions with polybasic regions of proteins. PI4Kα1 has been suggested to produce PI4P at the plasma membrane, but how it is recruited to this compartment is unknown. Here, we pin-point the mechanism that tethers PI4Kα1 to the plasma membrane via a nanodomain-anchored scaffolding complex. We identified that PI4Kα1 is part of a complex composed of proteins from the NO-POLLEN-GERMINATION, EFR3-OF-PLANTS, and HYCCIN-CONTAINING families. Comprehensive knock-out and knock-down strategies revealed that subunits of the PI4Kα1 complex are essential for pollen, embryonic and post-embryonic development. We further found that the PI4Kα1 complex is immobilized in plasma membrane nanodomains. Using synthetic mis-targeting strategies, we demonstrate that a combination of lipid anchoring and scaffolding localizes PI4Kα1 to the plasma membrane, which is essential for its function. Together, this work opens new perspectives on the mechanisms and function of plasma membrane nanopatterning by lipid kinases.

The lipid composition of organelles acts as a landmark to define membrane identity and specify subcellular function. Phosphoinositides are anionic lipids acting in protein sorting and trafficking at thetrans-Golgi network (TGN). In animal cells, sphingolipids are known to control the turnover of phosphoinositides through lipid exchange mechanisms at endoplasmic reticulum/TGN contact sites. In this study, we discovered a completely new mechanism acting on sphingolipid-mediated phosphoinositides homeostasis at the TGN in plant cells. We used multi-approaches to show that a reduction of the acyl-chain length of sphingolipid results in increased level of phosphatidylinositol-4-phosphate (PI4P) at the TGN, independently from either lipid exchange induced by sphingolipid synthetic flux, or local PI4P synthesis. Instead, we found that sphingolipids mediate the consumption of PI4P through phosphoinositide-specific phospholipase C (PI-PLC) and this process impacts the sorting of the auxin efflux carrier PIN2 at the TGN. Together, our data identify a new mode of action of sphingolipids in lipid interplay at the TGN during protein sorting.

Each phosphoinositide (PI, also known as phosphatidylinositol phosphate, polyphosphoinositide, PtdInsP or PIP) species is partitioned in the endomembrane system and thereby contributes to the identity of membrane compartments. However, membranes are in constant flux within this system, which raises the questions of how the spatiotemporal pattern of phosphoinositides is established and maintained within the cell. Here, we review the general mechanisms by which phosphoinositides and membrane trafficking feedbacks on each other to regulate cellular patterning. We then use the specific examples of polarized trafficking, endosomal sorting and vacuolar biogenesis to illustrate these general concepts.

Anionic phospholipids represent only minor fraction of cell membranes lipids but they are critically important for many membrane-related processes, including membrane identity, charge, shape, the generation of second messengers, and the recruitment of peripheral proteins. The main anionic phospholipids of the plasma membrane are phosphoinositides phosphatidylinositol 4-phosphate (PI4P), phosphatidylinositol 4,5-bisphosphate (PI4,5P

International audience; Anionic phospholipids represent only minor fraction of cell membranes lipids but they are critically important for many membrane-related processes, including membrane identity, charge, shape, the generation of second messengers and the recruitment of peripheral proteins. The main anionic phospholipids of the plasma membrane are phosphoinositides phosphatidylinositol 4-phosphate (PI4P), phosphatidylinositol 4,5-bisphosphate (PI4,5P 2), phosphatidylserine (PS), and phosphatidic acid (PA). Recent insights in the understanding of the nature of protein-phospholipid interactions enabled the design of genetically-encoded fluorescent molecular probes that can interact with various phospholipids in a specific manner allowing their imaging in live cells. Here, we describe the use of transiently transformed plant cells to study phospholipid-dependent membrane recruitment.

Membrane surface charge is critical for the transient, yet specific recruitment of proteins with polybasic regions to certain organelles. In all eukaryotes, the plasma membrane (PM) is the most electronegative compartment of the cell, which specifies its identity. As such, membrane electrostatics is a central parameter in signaling, intracellular trafficking and polarity. Here, we explore which are the lipids that control membrane electrostatics using plants as a model. We show that phosphatidic acidic (PA), phosphatidylserine (PS) and phosphatidylinositol-4-phosphate (PI4P) are separately required to generate the electrostatic signature of the plant PM. In addition, we reveal the existence of an electrostatic territory that is organized as a gradient along the endocytic pathway and is controlled by PS/PI4P combination. Altogether, we propose that combinatorial lipid composition of the cytosolic leaflet of cellular organelles not only defines the plant electrostatic territory but also distinguishes different compartments within this territory by specifying their varying surface charges.