TY - JOUR AB - Cell polarity is crucial for the coordinated development of all multicellular organisms. In plants, this is exemplified by the PIN-FORMED (PIN) efflux carriers of the phytohormone auxin: The polar subcellular localization of the PINs is instructive to the directional intercellular auxin transport, and thus to a plethora of auxin-regulated growth and developmental processes. Despite its importance, the regulation of PIN polar subcellular localization remains poorly understood. Here, we have employed advanced live-cell imaging techniques to study the roles of microtubules and actin microfilaments in the establishment of apical polar localization of PIN2 in the epidermis of the Arabidopsis root meristem. We report that apical PIN2 polarity requires neither intact actin microfilaments nor microtubules, suggesting that the primary spatial cue for polar PIN distribution is likely independent of cytoskeleton-guided endomembrane trafficking. AU - Glanc, Matous AU - Fendrych, Matyas AU - Friml, Jiří ID - 6611 IS - 6 JF - Biomolecules TI - PIN2 polarity establishment in arabidopsis in the absence of an intact cytoskeleton VL - 9 ER - TY - JOUR AB - An important adaptation during colonization of land by plants is gravitropic growth of roots, which enabled roots to reach water and nutrients, and firmly anchor plants in the ground. Here we provide insights into the evolution of an efficient root gravitropic mechanism in the seed plants. Architectural innovation, with gravity perception constrained in the root tips along with a shootward transport route for the phytohormone auxin, appeared only upon the emergence of seed plants. Interspecies complementation and protein domain swapping revealed functional innovations within the PIN family of auxin transporters leading to the evolution of gravitropism-specific PINs. The unique apical/shootward subcellular localization of PIN proteins is the major evolutionary innovation that connected the anatomically separated sites of gravity perception and growth response via the mobile auxin signal. We conclude that the crucial anatomical and functional components emerged hand-in-hand to facilitate the evolution of fast gravitropic response, which is one of the major adaptations of seed plants to dry land. AU - Zhang, Yuzhou AU - Xiao, G AU - Wang, X AU - Zhang, Xixi AU - Friml, Jiří ID - 6778 JF - Nature Communications SN - 2041-1723 TI - Evolution of fast root gravitropism in seed plants VL - 10 ER - TY - JOUR AB - Plants have a remarkable capacity to adjust their growth and development to elevated ambient temperatures. Increased elongation growth of roots, hypocotyls and petioles in warm temperatures are hallmarks of seedling thermomorphogenesis. In the last decade, significant progress has been made to identify the molecular signaling components regulating these growth responses. Increased ambient temperature utilizes diverse components of the light sensing and signal transduction network to trigger growth adjustments. However, it remains unknown whether temperature sensing and responses are universal processes that occur uniformly in all plant organs. Alternatively, temperature sensing may be confined to specific tissues or organs, which would require a systemic signal that mediates responses in distal parts of the plant. Here we show that Arabidopsis (Arabidopsis thaliana) seedlings show organ-specific transcriptome responses to elevated temperatures, and that thermomorphogenesis involves both autonomous and organ-interdependent temperature sensing and signaling. Seedling roots can sense and respond to temperature in a shoot-independent manner, whereas shoot temperature responses require both local and systemic processes. The induction of cell elongation in hypocotyls requires temperature sensing in cotyledons, followed by generation of a mobile auxin signal. Subsequently, auxin travels to the hypocotyl where it triggers local brassinosteroid-induced cell elongation in seedling stems, which depends upon a distinct, permissive temperature sensor in the hypocotyl. AU - Bellstaedt, Julia AU - Trenner, Jana AU - Lippmann, Rebecca AU - Poeschl, Yvonne AU - Zhang, Xixi AU - Friml, Jiří AU - Quint, Marcel AU - Delker, Carolin ID - 6366 IS - 2 JF - Plant Physiology SN - 0032-0889 TI - A mobile auxin signal connects temperature sensing in cotyledons with growth responses in hypocotyls VL - 180 ER - TY - JOUR AB - The plant hormone auxin has crucial roles in almost all aspects of plant growth and development. Concentrations of auxin vary across different tissues, mediating distinct developmental outcomes and contributing to the functional diversity of auxin. However, the mechanisms that underlie these activities are poorly understood. Here we identify an auxin signalling mechanism, which acts in parallel to the canonical auxin pathway based on the transport inhibitor response1 (TIR1) and other auxin receptor F-box (AFB) family proteins (TIR1/AFB receptors)1,2, that translates levels of cellular auxin to mediate differential growth during apical-hook development. This signalling mechanism operates at the concave side of the apical hook, and involves auxin-mediated C-terminal cleavage of transmembrane kinase 1 (TMK1). The cytosolic and nucleus-translocated C terminus of TMK1 specifically interacts with and phosphorylates two non-canonical transcriptional repressors of the auxin or indole-3-acetic acid (Aux/IAA) family (IAA32 and IAA34), thereby regulating ARF transcription factors. In contrast to the degradation of Aux/IAA transcriptional repressors in the canonical pathway, the newly identified mechanism stabilizes the non-canonical IAA32 and IAA34 transcriptional repressors to regulate gene expression and ultimately inhibit growth. The auxin–TMK1 signalling pathway originates at the cell surface, is triggered by high levels of auxin and shares a partially overlapping set of transcription factors with the TIR1/AFB signalling pathway. This allows distinct interpretations of different concentrations of cellular auxin, and thus enables this versatile signalling molecule to mediate complex developmental outcomes. AU - Cao, Min AU - Chen, Rong AU - Li, Pan AU - Yu, Yongqiang AU - Zheng, Rui AU - Ge, Danfeng AU - Zheng, Wei AU - Wang, Xuhui AU - Gu, Yangtao AU - Gelová, Zuzana AU - Friml, Jiří AU - Zhang, Heng AU - Liu, Renyi AU - He, Jun AU - Xu, Tongda ID - 6259 JF - Nature SN - 0028-0836 TI - TMK1-mediated auxin signalling regulates differential growth of the apical hook VL - 568 ER - TY - JOUR AB - PIN-FORMED (PIN) transporters mediate directional, intercellular movement of the phytohormone auxin in land plants. To elucidate the evolutionary origins of this developmentally crucial mechanism, we analysed the single PIN homologue of a simple green alga Klebsormidium flaccidum. KfPIN functions as a plasma membrane-localized auxin exporter in land plants and heterologous models. While its role in algae remains unclear, PIN-driven auxin export is probably an ancient and conserved trait within streptophytes. AU - Skokan, Roman AU - Medvecká, Eva AU - Viaene, Tom AU - Vosolsobě, Stanislav AU - Zwiewka, Marta AU - Müller, Karel AU - Skůpa, Petr AU - Karady, Michal AU - Zhang, Yuzhou AU - Janacek, Dorina P. AU - Hammes, Ulrich Z. AU - Ljung, Karin AU - Nodzyński, Tomasz AU - Petrášek, Jan AU - Friml, Jiří ID - 7106 IS - 11 JF - Nature Plants SN - 2055-0278 TI - PIN-driven auxin transport emerged early in streptophyte evolution VL - 5 ER - TY - JOUR AB - Roots grow downwards parallel to the gravity vector, to anchor a plant in soil and acquire water and nutrients, using a gravitropic mechanism dependent on the asymmetric distribution of the phytohormone auxin. Recently, Chang et al. demonstrate that asymmetric distribution of another phytohormone, cytokinin, directs root growth towards higher water content. AU - Sinclair, Scott A AU - Friml, Jiří ID - 7143 JF - Cell Research SN - 1001-0602 TI - Defying gravity: a plant's quest for moisture VL - 29 ER - TY - JOUR AB - During infection pathogens secrete small molecules, termed effectors, to manipulate and control the interaction with their specific hosts. Both the pathogen and the plant are under high selective pressure to rapidly adapt and co-evolve in what is usually referred to as molecular arms race. Components of the host’s immune system form a network that processes information about molecules with a foreign origin and damage-associated signals, integrating them with developmental and abiotic cues to adapt the plant’s responses. Both in the case of nucleotide-binding leucine-rich repeat receptors and leucine-rich repeat receptor kinases interaction networks have been extensively characterized. However, little is known on whether pathogenic effectors form complexes to overcome plant immunity and promote disease. Ustilago maydis, a biotrophic fungal pathogen that infects maize plants, produces effectors that target hubs in the immune network of the host cell. Here we assess the capability of U. maydis effector candidates to interact with each other, which may play a crucial role during the infection process. Using a systematic yeast-two-hybrid approach and based on a preliminary pooled screen, we selected 63 putative effectors for one-on-one matings with a library of nearly 300 effector candidates. We found that 126 of these effector candidates interacted either with themselves or other predicted effectors. Although the functional relevance of the observed interactions remains elusive, we propose that the observed abundance in complex formation between effectors adds an additional level of complexity to effector research and should be taken into consideration when studying effector evolution and function. Based on this fundamental finding, we suggest various scenarios which could evolutionarily drive the formation and stabilization of an effector interactome. AU - Alcântara, André AU - Bosch, Jason AU - Nazari, Fahimeh AU - Hoffmann, Gesa AU - Gallei, Michelle C AU - Uhse, Simon AU - Darino, Martin A. AU - Olukayode, Toluwase AU - Reumann, Daniel AU - Baggaley, Laura AU - Djamei, Armin ID - 7182 IS - 11 JF - Frontiers in Plant Science TI - Systematic Y2H screening reveals extensive effector-complex formation VL - 10 ER - TY - JOUR AB - Clathrin-mediated endocytosis (CME) is a highly conserved and essential cellular process in eukaryotic cells, but its dynamic and vital nature makes it challenging to study using classical genetics tools. In contrast, although small molecules can acutely and reversibly perturb CME, the few chemical CME inhibitors that have been applied to plants are either ineffective or show undesirable side effects. Here, we identify the previously described endosidin9 (ES9) as an inhibitor of clathrin heavy chain (CHC) function in both Arabidopsis and human cells through affinity-based target isolation, in vitro binding studies and X-ray crystallography. Moreover, we present a chemically improved ES9 analog, ES9-17, which lacks the undesirable side effects of ES9 while retaining the ability to target CHC. ES9 and ES9-17 have expanded the chemical toolbox used to probe CHC function, and present chemical scaffolds for further design of more specific and potent CHC inhibitors across different systems. AU - Dejonghe, Wim AU - Sharma, Isha AU - Denoo, Bram AU - De Munck, Steven AU - Lu, Qing AU - Mishev, Kiril AU - Bulut, Haydar AU - Mylle, Evelien AU - De Rycke, Riet AU - Vasileva, Mina K AU - Savatin, Daniel V. AU - Nerinckx, Wim AU - Staes, An AU - Drozdzecki, Andrzej AU - Audenaert, Dominique AU - Yperman, Klaas AU - Madder, Annemieke AU - Friml, Jiří AU - Van Damme, Daniël AU - Gevaert, Kris AU - Haucke, Volker AU - Savvides, Savvas N. AU - Winne, Johan AU - Russinova, Eugenia ID - 6377 IS - 6 JF - Nature Chemical Biology SN - 15524450 TI - Disruption of endocytosis through chemical inhibition of clathrin heavy chain function VL - 15 ER - TY - THES AB - The development and growth of Arabidopsis thaliana is regulated by a combination of genetic programing and also by the environmental influences. An important role in these processes play the phytohormones and among them, auxin is crucial as it controls many important functions. It is transported through the whole plant body by creating local and temporal concentration maxima and minima, which have an impact on the cell status, tissue and organ identity. Auxin has the property to undergo a directional and finely regulated cell-to-cell transport, which is enabled by the transport proteins, localized on the plasma membrane. An important role in this process have the PIN auxin efflux proteins, which have an asymmetric/polar subcellular localization and determine the directionality of the auxin transport. During the last years, there were significant advances in understanding how the trafficking molecular machineries function, including studies on molecular interactions, function, subcellular localization and intracellular distribution. However, there is still a lack of detailed characterization on the steps of endocytosis, exocytosis, endocytic recycling and degradation. Due to this fact, I focused on the identification of novel trafficking factors and better characterization of the intracellular trafficking pathways. My PhD thesis consists of an introductory chapter, three experimental chapters, a chapter containing general discussion, conclusions and perspectives and also an appendix chapter with published collaborative papers. The first chapter is separated in two different parts: I start by a general introduction to auxin biology and then I introduce the trafficking pathways in the model plant Arabidopsis thaliana. Then, I explain also the phosphorylation-signals for polar targeting and also the roles of the phytohormone strigolactone. The second chapter includes the characterization of bar1/sacsin mutant, which was identified in a forward genetic screen for novel trafficking components in Arabidopsis thaliana, where by the implementation of an EMS-treated pPIN1::PIN1-GFP marker line and by using the established inhibitor of ARF-GEFs, Brefeldin A (BFA) as a tool to study trafficking processes, we identified a novel factor, which is mediating the adaptation of the plant cell to ARF-GEF inhibition. The mutation is in a previously uncharacterized gene, encoding a very big protein that we, based on its homologies, called SACSIN with domains suggesting roles as a molecular chaperon or as a component of the ubiquitin-proteasome system. Our physiology and imaging studies revealed that SACSIN is a crucial plant cell component of the adaptation to the ARF-GEF inhibition. The third chapter includes six subchapters, where I focus on the role of the phytohormone strigolactone, which interferes with auxin feedback on PIN internalization. Strigolactone moderates the polar auxin transport by increasing the internalization of the PIN auxin efflux carriers, which reduces the canalization related growth responses. In addition, I also studied the role of phosphorylation in the strigolactone regulation of auxin feedback on PIN internalization. In this chapter I also present my results on the MAX2-dependence of strigolactone-mediated root growth inhibition and I also share my results on the auxin metabolomics profiling after application of GR24. In the fourth chapter I studied the effect of two small molecules ES-9 and ES9-17, which were identified from a collection of small molecules with the property to impair the clathrin-mediated endocytosis. In the fifth chapter, I discuss all my observations and experimental findings and suggest alternative hypothesis to interpret my results. In the appendix there are three collaborative published projects. In the first, I participated in the characterization of the role of ES9 as a small molecule, which is inhibitor of clathrin- mediated endocytosis in different model organisms. In the second paper, I contributed to the characterization of another small molecule ES9-17, which is a non-protonophoric analog of ES9 and also impairs the clathrin-mediated endocytosis not only in plant cells, but also in mammalian HeLa cells. Last but not least, I also attach another paper, where I tried to establish the grafting method as a technique in our lab to study canalization related processes. AU - Vasileva, Mina K ID - 7172 TI - Molecular mechanisms of endomembrane trafficking in Arabidopsis thaliana ER - TY - JOUR AB - Plasmodesmata (PD) are plant-specific membrane-lined channels that create cytoplasmic and membrane continuities between adjacent cells, thereby facilitating cell–cell communication and virus movement. Plant cells have evolved diverse mechanisms to regulate PD plasticity in response to numerous environmental stimuli. In particular, during defense against plant pathogens, the defense hormone, salicylic acid (SA), plays a crucial role in the regulation of PD permeability in a callose-dependent manner. Here, we uncover a mechanism by which plants restrict the spreading of virus and PD cargoes using SA signaling by increasing lipid order and closure of PD. We showed that exogenous SA application triggered the compartmentalization of lipid raft nanodomains through a modulation of the lipid raft-regulatory protein, Remorin (REM). Genetic studies, superresolution imaging, and transmission electron microscopy observation together demonstrated that Arabidopsis REM1.2 and REM1.3 are crucial for plasma membrane nanodomain assembly to control PD aperture and functionality. In addition, we also found that a 14-3-3 epsilon protein modulates REM clustering and membrane nanodomain compartmentalization through its direct interaction with REM proteins. This study unveils a molecular mechanism by which the key plant defense hormone, SA, triggers membrane lipid nanodomain reorganization, thereby regulating PD closure to impede virus spreading. AU - Huang, D AU - Sun, Y AU - Ma, Z AU - Ke, M AU - Cui, Y AU - Chen, Z AU - Chen, C AU - Ji, C AU - Tran, TM AU - Yang, L AU - Lam, SM AU - Han, Y AU - Shu, G AU - Friml, Jiří AU - Miao, Y AU - Jiang, L AU - Chen, X ID - 6999 IS - 42 JF - Proceedings of the National Academy of Sciences of the United States of America SN - 0027-8424 TI - Salicylic acid-mediated plasmodesmal closure via Remorin-dependent lipid organization VL - 116 ER -