Adaptor protein complex interaction map in Arabidopsis identifies P34 as a common stability regulator.

Adaptor protein (AP) complexes are evolutionarily conserved vesicle transport regulators that recruit coat proteins, membrane cargoes and coated vesicle accessory proteins. As in plants endocytic and post-Golgi trafficking intersect at the trans-Golgi network, unique mechanisms for sorting cargoes of overlapping vesicular routes are anticipated. The plant AP complexes are part of the sorting machinery, but despite some functional information, their cargoes, accessory proteins and regulation remain largely unknown. Here, by means of various proteomics approaches, we generated the overall interactome of the five AP and the TPLATE complexes in Arabidopsis thaliana. The interactome converged on a number of hub proteins, including the thus far unknown adaptin binding-like protein, designated P34. P34 interacted with the clathrin-associated AP complexes, controlled their stability and, subsequently, influenced clathrin-mediated endocytosis and various post-Golgi trafficking routes. Altogether, the AP interactome network offers substantial resources for further discoveries of unknown endomembrane trafficking regulators in plant cells.

Halotropism requires phospholipase Dζ1-mediated modulation of cellular polarity of auxin transport carriers.

Endocytosis and relocalization of auxin carriers represent important mechanisms for adaptive plant growth and developmental responses. Both root gravitropism and halotropism have been shown to be dependent on relocalization of auxin transporters. Following their homology to mammalian phospholipase Ds (PLDs), plant PLDζ-type enzymes are likely candidates to regulate auxin carrier endocytosis. We investigated root tropic responses for an Arabidopsis pldζ1-KO mutant and its effect on the dynamics of two auxin transporters during salt stress, that is, PIN2 and AUX1. We found altered root growth and halotropic and gravitropic responses in the absence of PLDζ1 and report a role for PLDζ1 in the polar localization of PIN2. Additionally, irrespective of the genetic background, salt stress induced changes in AUX1 polarity. Utilizing our previous computational model, we found that these novel salt-induced AUX1 changes contribute to halotropic auxin asymmetry. We also report the formation of "osmotic stress-induced membrane structures." These large membrane structures are formed at the plasma membrane shortly after NaCl or sorbitol treatment and have a prolonged presence in a pldζ1 mutant. Taken together, these results show a crucial role for PLDζ1 in both ionic and osmotic stress-induced auxin carrier dynamics during salt stress.

Out of Shape During Stress: A Key Role for Auxin.

In most abiotic stress conditions, including salinity and water deficit, the developmental plasticity of the plant root is regulated by the phytohormone auxin. Changes in auxin concentration are often attributed to changes in shoot-derived long-distance auxin flow. However, recent evidence suggests important contributions by short-distance auxin transport from local storage and local auxin biosynthesis, conjugation, and oxidation during abiotic stress. We discuss here current knowledge on long-distance auxin transport in stress responses, and subsequently debate how short-distance auxin transport and indole-3-acetic acid (IAA) metabolism play a role in influencing eventual auxin accumulation and signaling patterns. Our analysis stresses the importance of considering all these components together and highlights the use of mathematical modeling for predictions of plant physiological responses.

Modeling halotropism: a key role for root tip architecture and reflux loop remodeling in redistributing auxin.

A key characteristic of plant development is its plasticity in response to various and dynamically changing environmental conditions. Tropisms contribute to this flexibility by allowing plant organs to grow from or towards environmental cues. Halotropism is a recently described tropism in which plant roots bend away from salt. During halotropism, as in most other tropisms, directional growth is generated through an asymmetric auxin distribution that generates differences in growth rate and hence induces bending. Here, we develop a detailed model of auxin transport in the Arabidopsis root tip and combine this with experiments to investigate the processes generating auxin asymmetry during halotropism. Our model points to the key role of root tip architecture in allowing the decrease in PIN2 at the salt-exposed side of the root to result in a re-routing of auxin to the opposite side. In addition, our model demonstrates how feedback of auxin on the auxin transporter AUX1 amplifies this auxin asymmetry, while a salt-induced transient increase in PIN1 levels increases the speed at which this occurs. Using AUX1-GFP imaging and pin1 mutants, we experimentally confirmed these model predictions, thus expanding our knowledge of the cellular basis of halotropism.

Phosphatidic acid binding proteins display differential binding as a function of membrane curvature stress and chemical properties.

Phosphatidic acid (PA) is a crucial membrane phospholipid involved in de novo lipid synthesis and numerous intracellular signaling cascades. The signaling function of PA is mediated by peripheral membrane proteins that specifically recognize PA. While numerous PA-binding proteins are known, much less is known about what drives specificity of PA-protein binding. Previously, we have described the ionization properties of PA, summarized in the electrostatic-hydrogen bond switch, as one aspect that drives the specific binding of PA by PA-binding proteins. Here we focus on membrane curvature stress induced by phosphatidylethanolamine and show that many PA-binding proteins display enhanced binding as a function of negative curvature stress. This result is corroborated by the observation that positive curvature stress, induced by lyso phosphatidylcholine, abolishes PA binding of target proteins. We show, for the first time, that a novel plant PA-binding protein, Arabidopsis Epsin-like Clathrin Adaptor 1 (ECA1) displays curvature-dependence in its binding to PA. Other established PA targets examined in this study include, the plant proteins TGD2, and PDK1, the yeast proteins Opi1 and Spo20, and, the mammalian protein Raf-1 kinase and the C2 domain of the mammalian phosphatidylserine binding protein Lact as control. Based on our observations, we propose that liposome binding assays are the preferred method to investigate lipid binding compared to the popular lipid overlay assays where membrane environment is lost. The use of complex lipid mixtures is important to elucidate further aspects of PA binding proteins.

Halotropism is a response of plant roots to avoid a saline environment.

Tropisms represent fascinating examples of how plants respond to environmental signals by adapting their growth and development. Here, a novel tropism is reported, halotropism, allowing plant seedlings to reduce their exposure to salinity by circumventing a saline environment. In response to a salt gradient, Arabidopsis, tomato, and sorghum roots were found to actively prioritize growth away from salinity above following the gravity axis. Directionality of this response is established by an active redistribution of the plant hormone auxin in the root tip, which is mediated by the PIN-FORMED 2 (PIN2) auxin efflux carrier. We show that salt-induced phospholipase D activity stimulates clathrin-mediated endocytosis of PIN2 at the side of the root facing the higher salt concentration. The intracellular relocalization of PIN2 allows for auxin redistribution and for the directional bending of the root away from the higher salt concentration. Our results thus identify a cellular pathway essential for the integration of environmental cues with auxin-regulated root growth that likely plays a key role in plant adaptative responses to salt stress.