plant cell biology
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plant cell biology
science about plant cell organelles and development
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Frontiers | Single Event Resolution of Plant Plasma Membrane Protein Endocytosis by TIRF Microscopy | Plant Science

Frontiers | Single Event Resolution of Plant Plasma Membrane Protein Endocytosis by TIRF Microscopy | Plant Science | plant cell biology | Scoop.it
Endocytosis is a key process in the internalization of extracellular materials and plasma membrane proteins, such as receptors and transporters, thereby controlling many aspects of cell signaling and cellular homeostasis. Endocytosis in plants has an essential role not only for basic cellular functions but also for growth and development, nutrient delivery, toxin avoidance, and pathogen defense. The precise mechanisms of endocytosis in plants remain quite elusive. The lack of direct visualization and examination of single events of endocytosis has greatly hampered our ability to precisely monitor the cell surface lifetime and the recruitment profile of proteins driving endocytosis or endocytosed cargos in plants. Here we discuss the necessity to systematically implement total internal reflection fluorescence microcopy (TIRF) in the Plant Cell Biology community and present reliable protocols for high spatial and temporal imaging of endocytosis in plants using clathrin-mediated endocytosis (CME) as a test case, since it represents the major route for internalization of cell-surface proteins in plants. We developed a robust method to directly visualize cell surface proteins using TIRF microscopy combined to a high throughput, automated and unbiased analysis pipeline to determine the temporal recruitment profile of proteins to single sites of endocytosis, using the departure of clathrin as a physiological reference for scission. Using this ‘departure assay’, we assessed the recruitment of two different AP-2 subunits, alpha and mu, to the sites of endocytosis and found that AP2A1 was recruited in concert with clathrin, while AP2M was not. This validated approach therefore offers a powerful solution to better characterize the plant endocytic machinery and the dynamics of one’s favorite cargo protein.
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Was always wondering when some one uses TIRF on plant cells. Cool thing for sure!
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Life behind the wall: sensing mechanical cues in plants

Life behind the wall: sensing mechanical cues in plants | plant cell biology | Scoop.it
There is increasing evidence that all cells sense mechanical forces in order to perform their functions. In animals, mechanotransduction has been studied during the establishment of cell polarity, fate, and division in single cells, and increasingly is studied in the context of a multicellular tissue. What about plant systems? Our goal in this review is to summarize what is known about the perception of mechanical cues in plants, and to provide a brief comparison with animals.
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How selective severing by katanin promotes order in the plant cortical microtubule array

National Academy of Sciences
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System-wide organization of actin cytoskeleton determines organelle transport in hypocotyl plant cells

National Academy of Sciences
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Improving brightness and photostability of green and red fluorescent proteins for live cell imaging and FRET reporting

Improving brightness and photostability of green and red fluorescent proteins for live cell imaging and FRET reporting | plant cell biology | Scoop.it
Many genetically encoded biosensors use Förster resonance energy transfer (FRET) to dynamically report biomolecular activities.
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new ruby3 sounds promising for long term live cell imaging!
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Photo-convertible fluorescent proteins as tools for fresh insights on subcellular interactions in plants - GRIFFITHS - 2016 - Journal of Microscopy - Wiley Online Library

Photo-convertible fluorescent proteins as tools for fresh insights on subcellular interactions in plants - GRIFFITHS - 2016 - Journal of Microscopy - Wiley Online Library | plant cell biology | Scoop.it
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Chloroplast movement behavior varies widely among species and does not correlate with high light stress tolerance - Springer

Chloroplast movement behavior varies widely among species and does not correlate with high light stress tolerance - Springer | plant cell biology | Scoop.it
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One-Step Agrobacterium Mediated Transformation of E... [PLoS One. 2012] - PubMed - NCBI

Advancement in plant research is becoming impaired by the fact that the transfer of multiple genes is difficult to achieve. Here we present a new binary vector for Agrobacterium tumefaciens mediated transformation, pHUGE-Red, in concert with a cloning strategy suited for the transfer of up to nine genes at once. This vector enables modular cloning of large DNA fragments by employing Gateway technology and contains DsRED1 as visual selection marker. Furthermore, an R/Rs inducible recombination system was included allowing subsequent removal of the selection markers in the newly generated transgenic plants. We show the successful use of pHUGE-Red by transferring eight genes essential for Medicago truncatula to establish a symbiosis with rhizobia bacteria as one 74 kb T-DNA into four non-leguminous species; strawberry, poplar, tomato and tobacco. We provide evidence that all transgenes are expressed in the root tissue of the non-legumes. Visual control during the transformation process and subsequent marker gene removal makes the pHUGE-Red vector an excellent tool for the efficient transfer of multiple genes.

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PLoS ONE: Use of Confocal Laser as Light Source Reveals Stomata-Autonomous Function

PLoS ONE: Use of Confocal Laser as Light Source Reveals Stomata-Autonomous Function | plant cell biology | Scoop.it

An article, which reminds us that imaging itself can influence our beloved plant cells!

"In most terrestrial plants, stomata open during the day to maximize the update of CO2 for photosynthesis, but they close at night to minimize water loss. Blue light, among several environmental factors, controls this process. Stomata response to diverse stimuli seems to be dictated by the behaviour of neighbour stomata creating leaf areas of coordinated response. Here individual stomata of Arabidopsis leaves were illuminated with a short blue-light pulse by focusing a confocal argon laser. Beautifully, the illuminated stomata open their pores, whereas their dark-adapted neighbours unexpectedly experience no change. This induction of individual stomata opening by low fluence rates of blue light was disrupted in the phototropin1 phototropin2 (phot1 phot2) double mutant, which exhibits insensitivity of stomatal movements in blue-illuminated epidermal strips. The irradiation of all epidermal cells making direct contact with a given stoma in both wild type and phot1 phot2 plants does not trigger its movement. These results unravel the stoma autonomous function in the blue light response and illuminate the implication of PHOT1 and/or PHOT2 in such response. The micro spatial heterogeneity that solar blue light suffers in partially shaded leaves under natural conditions highlights the physiological significance of the autonomous stomatal behaviour."

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Actin-dependent plastid movement is required for motive force generation in directional nuclear movement in plant

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This is a nice read about how cells rearrange according to changing light conditions. Plastids pull the nucleus into the shadow! How nice of them :)

 

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Schattat Lab's curator insight, June 2, 2014 4:45 AM

This is a nice read about how cells rearrange according to changing light conditions. Plastids pull the nucleus into the shadow! How nice of them :)

 

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Ubiquitin facilitates a quality-control pathway that removes damaged chloroplasts

Ubiquitin facilitates a quality-control pathway that removes damaged chloroplasts | plant cell biology | Scoop.it
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ScienceDirect

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Review about our current knowledge of connecting membranes to actin in plant cells.
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Spatiotemporal monitoring of Pseudomonas effectors via type III secretion using split fluorescent protein fragments

Spatiotemporal monitoring of Pseudomonas effectors via type III secretion using split fluorescent protein fragments | plant cell biology | Scoop.it
Pathogenic gram-negative bacteria cause serious diseases in animals and plants. These bacterial pathogens use the type III secretion system (T3SS) to deliver effector proteins into host cells; these effectors then localize to different subcellular compartments to attenuate immune responses by altering biological processes of the host cells. The fluorescent protein (FP)-based approach to monitor effectors secreted from bacteria into the host cells is not possible because the folded FP prevents effector delivery through the T3SS. Therefore, we optimized an improved variant of self-assembling split superfolder green fluorescent protein (sfGFPOPT) system to investigate the spatiotemporal dynamics of effectors delivered through bacterial T3SS into plant cells. In this system, effectors are fused to 11th β-strand of super-folder GFP (sfGFP11) and when delivered into plant cells expressing sfGFP1-10 β-strand (sfGFP1-10OPT), the two proteins reconstitute GFP fluorescence. We generated a number of Arabidopsis thaliana transgenic lines expressing sfGFP1-10OPT targeted to various subcellular compartments to facilitate localization of GFP11-tagged effectors delivered from bacteria. We demonstrate the efficacy of this system using Pseudomonas effectors AvrB and AvrRps4 in Nicotiana benthamiana and transgenic Arabidopsis plants. The versatile split sfGFPOPT system described here will facilitate a better understanding of bacterial invasion strategies used to evade plant immune responses.
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TipTracker: automatic tracking of diverse moving objects - AoBBlog

TipTracker: automatic tracking of diverse moving objects - AoBBlog | plant cell biology | Scoop.it
Guest post by Arif Ashraf We study plant roots by observing their developmental stages, physiological conditions and their cell biology. The advent of fluo
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Hydrogen peroxide protects plants against sun damage

Hydrogen peroxide protects plants against sun damage | plant cell biology | Scoop.it
Plants use hydrogen peroxide -- best known for use in bleach and hair treatments -- to control how their cells react to varying levels of light, new research shows.
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Actin-dependence of the chloroplast cold positioning response in the liverwort Marchantia polymorpha L.

Actin-dependence of the chloroplast cold positioning response in the liverwort Marchantia polymorpha L. | plant cell biology | Scoop.it
The subcellular positioning of chloroplasts can be changed by alterations in the environment such as light and temperature. For example, in leaf mesophyll cells, chloroplasts localize along anticlinal cell walls under high-intensity light, and along periclinal cell walls under low-intensity light. These types of positioning responses are involved in photosynthetic optimization. In light-mediated chloroplast positioning responses, chloroplasts move to the appropriate positions in an actin-dependent manner, although some exceptions also depend on microtubule. Even under low-intensity light, at low temperature (e.g., 5°C), chloroplasts localize along anticlinal cell walls; this phenomenon is termed chloroplast cold positioning. In this study, we analyzed whether chloroplast cold positioning is dependent on actin filaments and/or microtubules in the liverwort Marchantia polymorpha L. When liverwort cells were treated with drugs for the de-polymerization of actin filaments, chloroplast cold positioning was completely inhibited. In contrast, chloroplast cold positioning was not affected by treatment with a drug for the de-polymerization of microtubules. These observations indicate the actin-dependence of chloroplast cold positioning in M. polymorpha. Actin filaments during the chloroplast cold positioning response were visualized by using fluorescent probes based on fluorescent proteins in living liverwort cells, and thus, their behavior during the chloroplast cold positioning response was documented.
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Long-Term Growth of Moss in Microfluidic Devices Enables Subcellular Studies in Development1[OPEN]

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nice tool and will be very helpfull!
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Frontiers | ER bodies in plants of the Brassicales order: biogenesis and association with innate immunity | Plant Cell Biology

Frontiers | ER bodies in plants of the Brassicales order: biogenesis and association with innate immunity | Plant Cell Biology | plant cell biology | Scoop.it
“ The endoplasmic reticulum (ER) forms highly organised network structures composed of tubules and cisternae. Many plant species develop additional ER-derived structures, most of which are specific f...”
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Screening a cDNA library for protein-protein inte... [Plant Cell. 2012] - PubMed - NCBI

Plant Cell. 2012 May;24(5):1746-59. Epub 2012 May 22.

Screening a cDNA library for protein-protein interactions directly in planta.

Lee LY et al.

Screening cDNA libraries for genes encoding proteins that interact with a bait protein is usually performed in yeast. However, subcellular compartmentation and protein modification may differ in yeast and plant cells, resulting in misidentification of protein partners. We used bimolecular fluorescence complementation technology to screen a plant cDNA library against a bait protein directly in plants. As proof of concept, we used the N-terminal fragment of yellow fluorescent protein- or nVenus-tagged Agrobacterium tumefaciens VirE2 and VirD2 proteins and the C-terminal extension (CTE) domain of Arabidopsis thaliana telomerase reverse transcriptase as baits to screen an Arabidopsis cDNA library encoding proteins tagged with the C-terminal fragment of yellow fluorescent protein. A library of colonies representing ∼2 × 10(5) cDNAs was arrayed in 384-well plates. DNA was isolated from pools of 10 plates, individual plates, and individual rows and columns of the plates. Sequential screening of subsets of cDNAs in Arabidopsis leaf or tobacco (Nicotiana tabacum) Bright Yellow-2 protoplasts identified single cDNA clones encoding proteins that interact with either, or both, of the Agrobacterium bait proteins, or with CTE. T-DNA insertions in the genes represented by some cDNAs revealed five novel Arabidopsis proteins important for Agrobacterium-mediated plant transformation. We also used this cDNA library to confirm VirE2-interacting proteins in orchid (Phalaenopsis amabilis) flowers. Thus, this technology can be applied to several plant species.

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Plant Methods Protocol: Streamlined sub-protocols for floral-dip transformation and selection of transformants in Arabidopsis thaliana

Generating and identifying transformants is essential for many studies of gene function. In Arabidopsis thaliana, a revolutionary protocol termed floral dip is now the most widely used transformation method. Although robust, it involves a number of relatively time-consuming and laborious steps, including manipulating an Agrobacterium tumefaciens culture and aseptic procedures for the selection of plant lines harboring antibiotic-selection markers. Furthermore, where multiple transgenes are to be introduced, achieving this by sequential transformations over multiple generations adds significantly to the time required. To circumvent these bottlenecks, we have developed three streamlined sub-protocols. First, we find that A. thaliana can be transformed by dipping directly into an A. tumefaciens culture supplemented with surfactant, eliminating the need for media exchange to a buffered solution. Next, we illustrate that A. thaliana lines possessing a double-transformation event can be readily generated by simply by floral-dipping into a mixture of two A. tumefaciens cultures harboring distinct transformation vectors. Finally, we report an alternative method of transformant selection on chromatography sand that does not require surface sterilization of seeds. These sub-protocols, which can be used separately or in combination, save time and money, and reduce the possibility of contamination.

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Plant Image Analysis - Software

Plant Image Analysis - Software | plant cell biology | Scoop.it

Website presenting the available plant image analysis softwares. This page is an amazing collection of hand pickend image Analysis tools for plant research!

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