@inbook{10268, abstract = {The analysis of dynamic cellular processes such as plant cytokinesis stands and falls with live-cell time-lapse confocal imaging. Conventional approaches to time-lapse imaging of cell division in Arabidopsis root tips are tedious and have low throughput. Here, we describe a protocol for long-term time-lapse simultaneous imaging of multiple root tips on a vertical-stage confocal microscope with automated root tracking. We also provide modifications of the basic protocol to implement this imaging method in the analysis of genetic, pharmacological or laser ablation wounding-mediated experimental manipulations. Our method dramatically improves the efficiency of cell division time-lapse imaging by increasing the throughput, while reducing the person-hour requirements of such experiments.}, author = {Hörmayer, Lukas and Friml, Jiří and Glanc, Matous}, booktitle = {Plant Cell Division}, isbn = {978-1-0716-1743-4}, issn = {1940-6029}, pages = {105--114}, publisher = {Humana Press}, title = {{Automated time-lapse imaging and manipulation of cell divisions in Arabidopsis roots by vertical-stage confocal microscopy}}, doi = {10.1007/978-1-0716-1744-1_6}, volume = {2382}, year = {2021}, } @article{8582, abstract = {Cell and tissue polarization is fundamental for plant growth and morphogenesis. The polar, cellular localization of Arabidopsis PIN‐FORMED (PIN) proteins is crucial for their function in directional auxin transport. The clustering of PIN polar cargoes within the plasma membrane has been proposed to be important for the maintenance of their polar distribution. However, the more detailed features of PIN clusters and the cellular requirements of cargo clustering remain unclear. Here, we characterized PIN clusters in detail by means of multiple advanced microscopy and quantification methods, such as 3D quantitative imaging or freeze‐fracture replica labeling. The size and aggregation types of PIN clusters were determined by electron microscopy at the nanometer level at different polar domains and at different developmental stages, revealing a strong preference for clustering at the polar domains. Pharmacological and genetic studies revealed that PIN clusters depend on phosphoinositol pathways, cytoskeletal structures and specific cell‐wall components as well as connections between the cell wall and the plasma membrane. This study identifies the role of different cellular processes and structures in polar cargo clustering and provides initial mechanistic insight into the maintenance of polarity in plants and other systems.}, author = {Li, Hongjiang and von Wangenheim, Daniel and Zhang, Xixi and Tan, Shutang and Darwish-Miranda, Nasser and Naramoto, Satoshi and Wabnik, Krzysztof T and de Rycke, Riet and Kaufmann, Walter and Gütl, Daniel J and Tejos, Ricardo and Grones, Peter and Ke, Meiyu and Chen, Xu and Dettmer, Jan and Friml, Jiří}, issn = {14698137}, journal = {New Phytologist}, number = {1}, pages = {351--369}, publisher = {Wiley}, title = {{Cellular requirements for PIN polar cargo clustering in Arabidopsis thaliana}}, doi = {10.1111/nph.16887}, volume = {229}, year = {2021}, } @article{8606, abstract = {The leaf is a crucial organ evolved with remarkable morphological diversity to maximize plant photosynthesis. The leaf shape is a key trait that affects photosynthesis, flowering rates, disease resistance, and yield. Although many genes regulating leaf development have been identified in the past years, the precise regulatory architecture underlying the generation of diverse leaf shapes remains to be elucidated. We used cotton as a reference model to probe the genetic framework underlying divergent leaf forms. Comparative transcriptome analysis revealed that the GhARF16‐1 and GhKNOX2‐1 genes might be potential regulators of leaf shape. We functionally characterized the auxin‐responsive factor ARF16‐1 acting upstream of GhKNOX2‐1 to determine leaf morphology in cotton. The transcription of GhARF16‐1 was significantly higher in lobed‐leaved cotton than in smooth‐leaved cotton. Furthermore, the overexpression of GhARF16‐1 led to the upregulation of GhKNOX2‐1 and resulted in more and deeper serrations in cotton leaves, similar to the leaf shape of cotton plants overexpressing GhKNOX2‐1. We found that GhARF16‐1 specifically bound to the promoter of GhKNOX2‐1 to induce its expression. The heterologous expression of GhARF16‐1 and GhKNOX2‐1 in Arabidopsis led to lobed and curly leaves, and a genetic analysis revealed that GhKNOX2‐1 is epistatic to GhARF16‐1 in Arabidopsis, suggesting that the GhARF16‐1 and GhKNOX2‐1 interaction paradigm also functions to regulate leaf shape in Arabidopsis. To our knowledge, our results uncover a novel mechanism by which auxin, through the key component ARF16‐1 and its downstream‐activated gene KNOX2‐1, determines leaf morphology in eudicots.}, author = {He, P and Zhang, Yuzhou and Li, H and Fu, X and Shang, H and Zou, C and Friml, Jiří and Xiao, G}, issn = {1467-7644}, journal = {Plant Biotechnology Journal}, number = {3}, pages = {548--562}, publisher = {Wiley}, title = {{GhARF16-1 modulates leaf development by transcriptionally regulating the GhKNOX2-1 gene in cotton}}, doi = {10.1111/pbi.13484}, volume = {19}, year = {2021}, } @article{8992, abstract = {The phytohormone auxin plays a central role in shaping plant growth and development. With decades of genetic and biochemical studies, numerous core molecular components and their networks, underlying auxin biosynthesis, transport, and signaling, have been identified. Notably, protein phosphorylation, catalyzed by kinases and oppositely hydrolyzed by phosphatases, has been emerging to be a crucial type of post-translational modification, regulating physiological and developmental auxin output at all levels. In this review, we comprehensively discuss earlier and recent advances in our understanding of genetics, biochemistry, and cell biology of the kinases and phosphatases participating in auxin action. We provide insights into the mechanisms by which reversible protein phosphorylation defines developmental auxin responses, discuss current challenges, and provide our perspectives on future directions involving the integration of the control of protein phosphorylation into the molecular auxin network.}, author = {Tan, Shutang and Luschnig, Christian and Friml, Jiří}, issn = {17529867}, journal = {Molecular Plant}, number = {1}, pages = {151--165}, publisher = {Elsevier}, title = {{Pho-view of auxin: Reversible protein phosphorylation in auxin biosynthesis, transport and signaling}}, doi = {10.1016/j.molp.2020.11.004}, volume = {14}, year = {2021}, } @article{8993, abstract = {N-1-naphthylphthalamic acid (NPA) is a key inhibitor of directional (polar) transport of the hormone auxin in plants. For decades, it has been a pivotal tool in elucidating the unique polar auxin transport-based processes underlying plant growth and development. Its exact mode of action has long been sought after and is still being debated, with prevailing mechanistic schemes describing only indirect connections between NPA and the main transporters responsible for directional transport, namely PIN auxin exporters. Here we present data supporting a model in which NPA associates with PINs in a more direct manner than hitherto postulated. We show that NPA inhibits PIN activity in a heterologous oocyte system and that expression of NPA-sensitive PINs in plant, yeast, and oocyte membranes leads to specific saturable NPA binding. We thus propose that PINs are a bona fide NPA target. This offers a straightforward molecular basis for NPA inhibition of PIN-dependent auxin transport and a logical parsimonious explanation for the known physiological effects of NPA on plant growth, as well as an alternative hypothesis to interpret past and future results. We also introduce PIN dimerization and describe an effect of NPA on this, suggesting that NPA binding could be exploited to gain insights into structural aspects of PINs related to their transport mechanism.}, author = {Abas, Lindy and Kolb, Martina and Stadlmann, Johannes and Janacek, Dorina P. and Lukic, Kristina and Schwechheimer, Claus and Sazanov, Leonid A and Mach, Lukas and Friml, Jiří and Hammes, Ulrich Z.}, issn = {10916490}, journal = {PNAS}, number = {1}, publisher = {National Academy of Sciences}, title = {{Naphthylphthalamic acid associates with and inhibits PIN auxin transporters}}, doi = {10.1073/pnas.2020857118}, volume = {118}, year = {2021}, } @article{9254, abstract = {Auxin is a key regulator of plant growth and development. Local auxin biosynthesis and intercellular transport generates regional gradients in the root that are instructive for processes such as specification of developmental zones that maintain root growth and tropic responses. Here we present a toolbox to study auxin-mediated root development that features: (i) the ability to control auxin synthesis with high spatio-temporal resolution and (ii) single-cell nucleus tracking and morphokinetic analysis infrastructure. Integration of these two features enables cutting-edge analysis of root development at single-cell resolution based on morphokinetic parameters under normal growth conditions and during cell-type-specific induction of auxin biosynthesis. We show directional auxin flow in the root and refine the contributions of key players in this process. In addition, we determine the quantitative kinetics of Arabidopsis root meristem skewing, which depends on local auxin gradients but does not require PIN2 and AUX1 auxin transporter activities. Beyond the mechanistic insights into root development, the tools developed here will enable biologists to study kinetics and morphology of various critical processes at the single cell-level in whole organisms.}, author = {Hu, Yangjie and Omary, Moutasem and Hu, Yun and Doron, Ohad and Hörmayer, Lukas and Chen, Qingguo and Megides, Or and Chekli, Ori and Ding, Zhaojun and Friml, Jiří and Zhao, Yunde and Tsarfaty, Ilan and Shani, Eilon}, issn = {20411723}, journal = {Nature Communications}, publisher = {Springer Nature}, title = {{Cell kinetics of auxin transport and activity in Arabidopsis root growth and skewing}}, doi = {10.1038/s41467-021-21802-3}, volume = {12}, year = {2021}, } @article{9443, abstract = {Endoplasmic reticulum–plasma membrane contact sites (ER–PM CS) play fundamental roles in all eukaryotic cells. Arabidopsis thaliana mutants lacking the ER–PM protein tether synaptotagmin1 (SYT1) exhibit decreased PM integrity under multiple abiotic stresses, such as freezing, high salt, osmotic stress, and mechanical damage. Here, we show that, together with SYT1, the stress-induced SYT3 is an ER–PM tether that also functions in maintaining PM integrity. The ER–PM CS localization of SYT1 and SYT3 is dependent on PM phosphatidylinositol-4-phosphate and is regulated by abiotic stress. Lipidomic analysis revealed that cold stress increased the accumulation of diacylglycerol at the PM in a syt1/3 double mutant relative to wild-type while the levels of most glycerolipid species remain unchanged. In addition, the SYT1-green fluorescent protein fusion preferentially binds diacylglycerol in vivo with little affinity for polar glycerolipids. Our work uncovers a SYT-dependent mechanism of stress adaptation counteracting the detrimental accumulation of diacylglycerol at the PM produced during episodes of abiotic stress.}, author = {Ruiz-Lopez, N and Pérez-Sancho, J and Esteban Del Valle, A and Haslam, RP and Vanneste, S and Catalá, R and Perea-Resa, C and Van Damme, D and García-Hernández, S and Albert, A and Vallarino, J and Lin, J and Friml, Jiří and Macho, AP and Salinas, J and Rosado, A and Napier, JA and Amorim-Silva, V and Botella, MA}, issn = {1532-298x}, journal = {Plant Cell}, number = {7}, pages = {2431--2453}, publisher = {American Society of Plant Biologists}, title = {{Synaptotagmins at the endoplasmic reticulum-plasma membrane contact sites maintain diacylglycerol homeostasis during abiotic stress}}, doi = {10.1093/plcell/koab122}, volume = {33}, year = {2021}, } @article{9657, abstract = {To overcome nitrogen deficiency, legume roots establish symbiotic interactions with nitrogen-fixing rhizobia that is fostered in specialized organs (nodules). Similar to other organs, nodule formation is determined by a local maximum of the phytohormone auxin at the primordium site. However, how auxin regulates nodule development remains poorly understood. Here, we found that in soybean, (Glycine max), dynamic auxin transport driven by PIN-FORMED (PIN) transporter GmPIN1 is involved in nodule primordium formation. GmPIN1 was specifically expressed in nodule primordium cells and GmPIN1 was polarly localized in these cells. Two nodulation regulators, (iso)flavonoids trigger expanded distribution of GmPIN1b to root cortical cells, and cytokinin rearranges GmPIN1b polarity. Gmpin1abc triple mutants generated with CRISPR-Cas9 showed impaired establishment of auxin maxima in nodule meristems and aberrant divisions in the nodule primordium cells. Moreover, overexpression of GmPIN1 suppressed nodule primordium initiation. GmPIN9d, an ortholog of Arabidopsis thaliana PIN2, acts together with GmPIN1 later in nodule development to acropetally transport auxin in vascular bundles, fine-tuning the auxin supply for nodule enlargement. Our findings reveal how PIN-dependent auxin transport modulates different aspects of soybean nodule development and suggest that establishment of auxin gradient is a prerequisite for the proper interaction between legumes and rhizobia.}, author = {Gao, Z and Chen, Z and Cui, Y and Ke, M and Xu, H and Xu, Q and Chen, J and Li, Y and Huang, L and Zhao, H and Huang, D and Mai, S and Xu, T and Liu, X and Li, S and Guan, Y and Yang, W and Friml, Jiří and Petrášek, J and Zhang, J and Chen, X}, issn = {1532-298x}, journal = {Plant Cell}, number = {9}, pages = {2981–3003}, publisher = {American Society of Plant Biologists}, title = {{GmPIN-dependent polar auxin transport is involved in soybean nodule development}}, doi = {10.1093/plcell/koab183}, volume = {33}, year = {2021}, } @article{9656, abstract = {Tropisms, growth responses to environmental stimuli such as light or gravity, are spectacular examples of adaptive plant development. The plant hormone auxin serves as a major coordinative signal. The PIN auxin exporters, through their dynamic polar subcellular localizations, redirect auxin fluxes in response to environmental stimuli and the resulting auxin gradients across organs underly differential cell elongation and bending. In this review, we discuss recent advances concerning regulations of PIN polarity during tropisms, focusing on PIN phosphorylation and trafficking. We also cover how environmental cues regulate PIN actions during tropisms, and a crucial role of auxin feedback on PIN polarity during bending termination. Finally, the interactions between different tropisms are reviewed to understand plant adaptive growth in the natural environment.}, author = {Han, Huibin and Adamowski, Maciek and Qi, Linlin and Alotaibi, SS and Friml, Jiří}, issn = {1469-8137}, journal = {New Phytologist}, number = {2}, pages = {510--522}, publisher = {Wiley}, title = {{PIN-mediated polar auxin transport regulations in plant tropic responses}}, doi = {10.1111/nph.17617}, volume = {232}, year = {2021}, } @article{9909, abstract = {Roots are composed of different root types and, in the dicotyledonous Arabidopsis, typically consist of a primary root that branches into lateral roots. Adventitious roots emerge from non-root tissue and are formed upon wounding or other types of abiotic stress. Here, we investigated adventitious root (AR) formation in Arabidopsis hypocotyls under conditions of altered abscisic acid (ABA) signaling. Exogenously applied ABA suppressed AR formation at 0.25 µM or higher doses. AR formation was less sensitive to the synthetic ABA analog pyrabactin (PB). However, PB was a more potent inhibitor at concentrations above 1 µM, suggesting that it was more selective in triggering a root inhibition response. Analysis of a series of phosphonamide and phosphonate pyrabactin analogs suggested that adventitious root formation and lateral root branching are differentially regulated by ABA signaling. ABA biosynthesis and signaling mutants affirmed a general inhibitory role of ABA and point to PYL1 and PYL2 as candidate ABA receptors that regulate AR inhibition.}, author = {Zeng, Yinwei and Verstraeten, Inge and Trinh, Hoang Khai and Heugebaert, Thomas and Stevens, Christian V. and Garcia-Maquilon, Irene and Rodriguez, Pedro L. and Vanneste, Steffen and Geelen, Danny}, issn = {20734425}, journal = {Genes}, number = {8}, publisher = {MDPI}, title = {{Arabidopsis hypocotyl adventitious root formation is suppressed by ABA signaling}}, doi = {10.3390/genes12081141}, volume = {12}, year = {2021}, }