Other Publications (7)
- Current Opinion in Plant Biology
- Journal of Plant Physiology
- BMC Plant Biology
- Microscopy and Microanalysis : the Official Journal of Microscopy Society of America, Microbeam Analysis Society, Microscopical Society of Canada
- Biochimica Et Biophysica Acta
- International Journal of Molecular Sciences
- Plant Cell Reports
Articles by Jan Petrášek in JoVE
Determination of Plasma Membrane Partitioning for Peripherally-associated Proteins Stanislav Vosolsobě1, Kateřina Schwarzerová1, Jan Petrášek1 1Department of Experimental Plant Biology, University of Science, Charles University Here, we present a protocol to perform a quantitative analysis of the level of plasma-membrane association for fluorescently-tagged peripherally-associated protein. The method is based on the computational decomposition of membrane and cytoplasmic component of signal observed in cells labeled with plasma membrane fluorescent marker.
Other articles by Jan Petrášek on PubMed
Actin and Microtubule Cytoskeleton Interactions Current Opinion in Plant Biology. | Pubmed ID: 19854097 Plant cytoskeleton consists of two major networks of protein polymers, actin microfilaments (AFs) and microtubules (MTs). These networks perform numerous functions that are essential for cell division and for maintaining the integrity of cytoplasm required for intracellular transport and cell shape. Besides the more or less indirect cooperation between AFs and MTs, their direct interactions through specific physically interacting proteins has been well described in yeast, nematodes, insect and animal cells. Recently, promising candidates for corresponding homologous proteins have been identified in plants, although there is still lack of functional evidence for these interactions. Here we summarize recent advances in our knowledge about the candidate proteins or protein complexes that interact with both AFs and MTs and their role in fundamental cellular and developmental processes.
Characterization of Transmembrane Auxin Transport in Arabidopsis Suspension-cultured Cells Journal of Plant Physiology. | Pubmed ID: 24594395 Polar auxin transport is a crucial process for control and coordination of plant development. Studies of auxin transport through plant tissues and organs showed that auxin is transported by a combination of phloem flow and the active, carrier-mediated cell-to-cell transport. Since plant organs and even tissues are too complex for determination of the kinetics of carrier-mediated auxin uptake and efflux on the cellular level, simplified models of cell suspension cultures are often used, and several tobacco cell lines have been established for auxin transport assays. However, there are very few data available on the specificity and kinetics of auxin transport across the plasma membrane for Arabidopsis thaliana suspension-cultured cells. In this report, the characteristics of carrier-mediated uptake (influx) and efflux for the native auxin indole-3-acetic acid and synthetic auxins, naphthalene-1-acetic and 2,4-dichlorophenoxyacetic acids (NAA and 2,4-D, respectively) in A. thaliana ecotype Landsberg erecta suspension-cultured cells (LE line) are provided. By auxin competition assays and inhibitor treatments, we show that, similarly to tobacco cells, uptake carriers have high affinity towards 2,4-D and that NAA is a good tool for studies of auxin efflux in LE cells. In contrast to tobacco cells, metabolic profiling showed that only a small proportion of NAA is metabolized in LE cells. These results show that the LE cell line is a useful experimental system for measurements of kinetics of auxin carriers on the cellular level that is complementary to tobacco cells.
NtGNL1a ARF-GEF Acts in Endocytosis in Tobacco Cells BMC Plant Biology. | Pubmed ID: 26541824 Processes of anterograde and retrograde membrane trafficking play an important role in cellular homeostasis and dynamic rearrangements of the plasma membrane (PM) in all eukaryotes. These processes depend on the activity of adenosine ribosylation factors (ARFs), a family of GTP-binding proteins and their guanine exchange factors (GEFs). However, knowledge on the function and specificity of individual ARF-GEFs for individual steps of membrane trafficking pathways is still limited in plants.
Determination of Dynamics of Plant Plasma Membrane Proteins with Fluorescence Recovery and Raster Image Correlation Spectroscopy Microscopy and Microanalysis : the Official Journal of Microscopy Society of America, Microbeam Analysis Society, Microscopical Society of Canada. | Pubmed ID: 27041337 A number of fluorescence microscopy techniques are described to study dynamics of fluorescently labeled proteins, lipids, nucleic acids, and whole organelles. However, for studies of plant plasma membrane (PM) proteins, the number of these techniques is still limited because of the high complexity of processes that determine the dynamics of PM proteins and the existence of cell wall. Here, we report on the usage of raster image correlation spectroscopy (RICS) for studies of integral PM proteins in suspension-cultured tobacco cells and show its potential in comparison with the more widely used fluorescence recovery after photobleaching method. For RICS, a set of microscopy images is obtained by single-photon confocal laser scanning microscopy (CLSM). Fluorescence fluctuations are subsequently correlated between individual pixels and the information on protein mobility are extracted using a model that considers processes generating the fluctuations such as diffusion and chemical binding reactions. As we show here using an example of two integral PM transporters of the plant hormone auxin, RICS uncovered their distinct short-distance lateral mobility within the PM that is dependent on cytoskeleton and sterol composition of the PM. RICS, which is routinely accessible on modern CLSM instruments, thus represents a valuable approach for studies of dynamics of PM proteins in plants.
Evolutionary Plasticity of Plasma Membrane Interaction in DREPP Family Proteins Biochimica Et Biophysica Acta. | Pubmed ID: 28108158 The plant-specific DREPP protein family comprises proteins that were shown to regulate the actin and microtubular cytoskeleton in a calcium-dependent manner. Our phylogenetic analysis showed that DREPPs first appeared in ferns and that DREPPs have a rapid and plastic evolutionary history in plants. Arabidopsis DREPP paralogues called AtMDP25/PCaP1 and AtMAP18/PCaP2 are N-myristoylated, which has been reported as a key factor in plasma membrane localization. Here we show that N-myristoylation is neither conserved nor ancestral for the DREPP family. Instead, by using confocal microscopy and a new method for quantitative evaluation of protein membrane localization, we show that DREPPs rely on two mechanisms ensuring their plasma membrane localization. These include N-myristoylation and electrostatic interaction of a polybasic amino acid cluster. We propose that various plasma membrane association mechanisms resulting from the evolutionary plasticity of DREPPs are important for refining plasma membrane interaction of these signalling proteins under various conditions and in various cells.
Evolutionary Conserved Cysteines Function As Cis-Acting Regulators of Arabidopsis PIN-FORMED 2 Distribution International Journal of Molecular Sciences. | Pubmed ID: 29109378 Coordination of plant development requires modulation of growth responses that are under control of the phytohormone auxin. PIN-FORMED plasma membrane proteins, involved in intercellular transport of the growth regulator, are key to the transmission of such auxin signals and subject to multilevel surveillance mechanisms, including reversible post-translational modifications. Apart from well-studied PIN protein modifications, namely phosphorylation and ubiquitylation, no further post-translational modifications have been described so far. Here, we focused on root-specific PIN2 and explored functional implications of two evolutionary conserved cysteines, by a combination of and molecular approaches. PIN2 sequence alignments and modeling predictions indicated that both cysteines are facing the cytoplasm and therefore would be accessible to redox status-controlled modifications. Notably, mutant alleles retained functionality, demonstrated by their ability to almost completely rescue defects of a null allele, whereas high resolution analysis of pin2 localization revealed increased intracellular accumulation, and altered protein distribution within plasma membrane micro-domains. The observed effects of cysteine replacements on root growth and PIN2 localization are consistent with a model in which redox status-dependent cysteine modifications participate in the regulation of PIN2 mobility, thereby fine-tuning polar auxin transport.
Silver Ions Increase Plasma Membrane Permeability Through Modulation of Intracellular Calcium Levels in Tobacco BY-2 Cells Plant Cell Reports. | Pubmed ID: 29502206 Silver ions increase plasma membrane permeability for water and small organic compounds through their stimulatory effect on plasma membrane calcium channels, with subsequent modulation of intracellular calcium levels and ion homeostasis. The action of silver ions at the plant plasma membrane is largely connected with the inhibition of ethylene signalling thanks to the ability of silver ion to replace the copper cofactor in the ethylene receptor. A link coupling the action of silver ions and cellular auxin efflux has been suggested earlier by their possible direct interaction with auxin efflux carriers or by influencing plasma membrane permeability. Using tobacco BY-2 cells, we demonstrate here that besides a dramatic increase of efflux of synthetic auxins 2,4-dichlorophenoxyacetic acid (2,4-D) and 1-naphthalene acetic acid (NAA), treatment with AgNO resulted in enhanced efflux of the cytokinin trans-zeatin (tZ) as well as the auxin structural analogues tryptophan (Trp) and benzoic acid (BA). The application of AgNO was accompanied by gradual water loss and plasmolysis. The observed effects were dependent on the availability of extracellular calcium ions (Ca) as shown by comparison of transport assays in Ca-rich and Ca-free buffers and upon treatment with inhibitors of plasma membrane Ca-permeable channels Al and ruthenium red, both abolishing the effect of AgNO. Confocal microscopy of Ca-sensitive fluorescence indicator Fluo-4FF, acetoxymethyl (AM) ester suggested that the extracellular Ca availability is necessary to trigger the response to silver ions and that the intracellular Ca pool alone is not sufficient for this effect. Altogether, our data suggest that in plant cells the effects of silver ions originate from the primal modification of the internal calcium levels, possibly by their interaction with Ca-permeable channels at the plasma membrane.