TY - JOUR AU - Friml, Jirí AU - Benfey, Philip AU - Benková, Eva AU - Bennett, Malcolm AU - Berleth, Thomas AU - Geldner, Niko AU - Grebe, Markus AU - Heisler, Marcus AU - Hejátko, Jan AU - Jürgens, Gerd AU - Laux, Thomas AU - Lindsey, Keith AU - Lukowitz, Wolfgang AU - Luschnig, Christian AU - Offringa, Remko AU - Scheres, Ben AU - Swarup, Ranjan AU - Torres Ruiz, Ramón AU - Weijers, Dolf AU - Zažímalová, Eva ID - 3005 IS - 1 JF - Trends in Plant Science TI - Apical-basal polarity: Why plant cells don't stand on their heads VL - 11 ER - TY - JOUR AB - Plants and some animals have a profound capacity to regenerate organs from adult tissues. Molecular mechanisms for regeneration have, however, been largely unexplored. Here we investigate a local regeneration response in Arabidopsis roots. Laser-induced wounding disrupts the flow of auxin-a cell-fate-instructive plant hormone-in root tips, and we demonstrate that resulting cell-fate changes require the PLETHORA, SHORTROOT, and SCARECROW transcription factors. These transcription factors regulate the expression and polar position of PIN auxin efflux-facilitating membrane proteins to reconstitute auxin transport in renewed root tips. Thus, a regeneration mechanism using embryonic root stem-cell patterning factors first responds to and subsequently stabilizes a new hormone distribution. AU - Xu, Jian AU - Hofhuis, Hugo AU - Heidstra, Renze AU - Sauer, Michael AU - Jirí Friml AU - Scheres, Ben ID - 3008 IS - 5759 JF - Science TI - A molecular framework for plant regeneration VL - 311 ER - TY - JOUR AU - Paciorek, Tomasz AU - Friml, Jirí ID - 3009 IS - 7 JF - Journal of Cell Science TI - Auxin signaling VL - 119 ER - TY - JOUR AB - Plant development is characterized by a profound ability to regenerate and form tissues with new axes of polarity. An unsolved question concerns how the position within a tissue and cues from neighboring cells are integrated to specify the polarity of individual cells. The canalization hypothesis proposes a feedback effect of the phytohormone auxin on the directionality of intercellular auxin flow as a means to polarize tissues. Here we identify a cellular and molecular mechanism for canalization. Local auxin application, wounding, or auxin accumulation during de novo organ formation lead to rearrangements in the subcellular polar localization of PIN auxin transport components. This auxin effect on PIN polarity is cell-specific, does not depend on PIN transcription, and involves the Aux/IAA-ARF (indole-3-acetic acid-auxin response factor) signaling pathway. Our data suggest that auxin acts as polarizing cue, which links individual cell polarity with tissue and organ polarity through control of PIN polar targeting. This feedback regulation provides a conceptual framework for polarization during multiple regenerative and patterning processes in plants. AU - Sauer, Michael AU - Balla, Jozef AU - Luschnig, Christian AU - Wiśniewska, Justyna AU - Reinöhl, Vilém AU - Friml, Jirí AU - Benková, Eva ID - 3016 IS - 20 JF - Genes and Development TI - Canalization of auxin flow by Aux/IAA-ARF-dependent feedback regulation of PIN polarity VL - 20 ER - TY - JOUR AB - The plant hormone auxin plays crucial roles in regulating plant growth development, including embryo and root patterning, organ formation, vascular tissue differentiation and growth responses to environmental stimuli. Asymmetric auxin distribution patterns have been observed within tissues, and these so-called auxin gradients change dynamically during different developmental processes. Most auxin is synthesized in the shoot and distributed directionally throughout the plant. This polar auxin transport is mediated by auxin influx and efflux facilitators, whose subcellular polar localizations guide the direction of auxin flow. The polar localization of PIN auxin efflux carriers changes in response to developmental and external cues in order to channel auxin flow in a regulated manner for organized growth. Auxin itself modulates the expression and subcellular localization of PIN proteins, contributing to a complex pattern of feedback regulation. Here we review the available information mainly from studies of a model plant, Arabidopsis thaliana, on the generation of auxin gradients, the regulation of polar auxin transport and further downstream cellular events. AU - Tanaka, Hirokazu AU - Dhonukshe, Pankaj AU - Brewer, Philip AU - Friml, Jirí ID - 3017 IS - 23 JF - Cellular and Molecular Life Sciences TI - Spatiotemporal asymmetric auxin distribution: A means to coordinate plant development VL - 63 ER - TY - JOUR AB - The directional flow of the plant hormone auxin mediates multiple developmental processes, including patterning and tropisms. Apical and basal plasma membrane localization of AUXIN-RESISTANT1 (AUX1) and PIN-FORMED1 (PIN1) auxin transport components underpins the directionality of intercellular auxin flow in Arabidopsis thaliana roots. Here, we examined the mechanism of polar trafficking of AUX1. Real-time live cell analysis along with subcellular markers revealed that AUX1 resides at the apical plasma membrane of protophloem cells and at highly dynamic subpopulations of Golgi apparatus and endosomes in all cell types. Plasma membrane and intracellular pools of AUX1 are interconnected by actin-dependent constitutive trafficking, which is not sensitive to the vesicle trafficking inhibitor brefeldin A. AUX1 subcellular dynamics are not influenced by the auxin influx inhibitor NOA but are blocked by the auxin efflux inhibitors TIBA and PBA. Furthermore, auxin transport inhibitors and interference with the sterol composition of membranes disrupt polar AUX1 distribution at the plasma membrane. Compared with PIN1 trafficking, AUX1 dynamics display different sensitivities to trafficking inhibitors and are independent of the endosomal trafficking regulator ARF GEF GNOM. Hence, AUX1 uses a novel trafficking pathway in plants that is distinct from PIN trafficking, providing an additional mechanism for the fine regulation of auxin transport. AU - Kleine-Vehn, Jürgen AU - Dhonukshe, Pankaj AU - Swarup, Ranjan AU - Bennett, Malcolm AU - Jirí Friml ID - 3018 IS - 11 JF - Plant Cell TI - Subcellular trafficking of the Arabidopsis auxin influx carrier AUX1 uses a novel pathway distinct from PIN1 VL - 18 ER - TY - JOUR AB - High throughput microarray transcription analyses provide us with the expression profiles for large amounts of plant genes. However, their tissue and cellular resolution is limited. Thus, for detailed functional analysis, it is still necessary to examine the expression pattern of selected candidate genes at a cellular level. Here, we present an in situ mRNA hybridization method that is routinely used for the analysis of plant gene expression patterns. The protocol is optimized for whole mount mRNA localizations in Arabidopsis seedling tissues including embryos, roots, hypocotyls and young primary leaves. It can also be used for comparable tissues in other species. Part of the protocol can also be automated and performed by a liquid handling robot. Here we present a detailed protocol, recommended controls and troubleshooting, along with examples of several applications. The total time to carry out the entire procedure is ∼7 d, depending on the tissue used. AU - Hejátko, Jan AU - Blilou, Ikram AU - Brewer, Philip B AU - Jirí Friml AU - Scheres, Ben AU - Eva Benková ID - 3020 IS - 4 JF - Nature Protocols TI - In situ hybridization technique for mRNA detection in whole mount Arabidopsis samples VL - 1 ER - TY - JOUR AB - As the field of plant molecular biology is swiftly advancing, a need has been created for methods that allow rapid and reliable in situ localization of proteins in plant cells. Here we describe a whole-mount 'immunolocalization' technique for various plant tissues, including roots, hypocotyls, cotyledons, young primary leaves and embryos of Arabidopsis thaliana and other species. The detailed protocol, recommended controls and troubleshooting are presented, along with examples of applications. The protocol consists of five main procedures: tissue fixation, tissue permeation, blocking, primary and secondary antibody incubation. Notably, the first procedure (tissue fixation) includes several steps (4-12) that are absolutely necessary for protein localization in hypocotyls, cotyledons and young primary leaves but should be omitted for other tissues. The protocol is usually done in 3 days, but could also be completed in 2 days. AU - Sauer, Michael AU - Paciorek, Tomasz AU - Eva Benková AU - Jirí Friml ID - 3015 IS - 1 JF - Nature Protocols TI - Immunocytochemical techniques for whole mount in situ protein localization in plants VL - 1 ER - TY - JOUR AB - There is a growing demand for methods that allow rapid and reliable in situ localization of proteins in plant cells. The immunocytochemistry protocol presented here can be used routinely to observe protein localization patterns in tissue sections of various plant species. This protocol is especially suitable for plant species with more-complex tissue architecture (such as maize, Zea mays), which makes it difficult to use an easier whole-mount procedure for protein localization. To facilitate the antibody-antigen reaction, it is necessary to include a wax-embedding and tissue-sectioning step. The protocol consists of the following procedures: chemical fixation of tissue, dehydration, wax embedding, sectioning, dewaxing, rehydration, blocking and antibody incubation. The detailed protocol, recommended controls and troubleshooting are presented here, along with examples of applications. AU - Paciorek, Tomasz AU - Sauer, Michael AU - Balla, Jozef AU - Wiśniewska, Justyna AU - Jirí Friml ID - 3013 IS - 1 JF - Nature Protocols TI - Immunocytochemical technique for protein localization in sections of plant tissues VL - 1 ER - TY - JOUR AB - Plant biology is currently confronted with an overflow of expression profile data provided by high-throughput microarray transcription analyses. However, the tissue and cellular resolution of these techniques is limited. Thus, it is still necessary to examine the expression pattern of selected candidate genes at a cellular level. Here we present an in situ mRNA hybridization method that is routinely used in the analysis of gene expression patterns. The protocol is optimized for mRNA localizations in sectioned tissue of Arabidopsis seedlings including embryos, roots, hypocotyls, young primary leaves and flowers. The detailed protocol, recommended controls and troubleshooting are presented along with examples of application. The total time for the process is 10 days. AU - Brewer, Philip B AU - Heisler, Marcus G AU - Hejátko, Jan AU - Jirí Friml AU - Eva Benková ID - 3014 IS - 3 JF - Nature Protocols TI - In situ hybridization for mRNA detection in Arabidopsis tissue sections VL - 1 ER -