Cell Perception Research Articles

Never Walk Alone: Clathrin-coated Vesicle (CCV) Components in Plant Immunity

The plasma's protein makeup at the host-pathogen interaction membrane (PM) has significant effects on a plant cell's perception of and reacts to microbiological infections that invade. The capacity of a plant to adjust Its PM composition is essential for controlling the intensity, length, and immunological response integration. One method by which plant cells reconfigure the vesicular trafficking on their cell surface, encompassing secretion and endocytosis. During these trafficking procedures, cargo proteins (such as transporters, receptors for pattern recognition, and other proteins with immunological activities) by tiny, membrane-bound vesicles to or from the PM. Vesicles covered with clathrin (CCVs) that develop at the PM and trans-Golgi membranes Early endosomes and networks have become the most common vesicle form in the control of plant defense mechanisms.

Molecular biology of the cell

Molecular biology of the cell

Membrane dynamics in physiology and disease

The composition and dynamics of cellular membranes determine intra- and extracellular signaling events, and modulate a variety of cellular states including proliferation, survival, migration and more. Eight minireviews covering some aspects of the ESF EuroMEMBRANE programme and conference elucidate phosphoinositide signaling, membrane sorting, topography, lipid dynamics modeling, and mass spectroscopy methods to enlarge the known lipidome. Membranes define the identity of cells: they are crucial for a cell's perception of the environment, they regulate cellular homeostasis, and they function as hubs for extra- and intracellular signals. Countless lipids have been identified, but how they function in various cell organelles is still poorly defined. Dynamic processes such as membrane fusion, fission, aging and elimination are being elucidated, but are not fully understood. We know that localized lipid–protein and lipid–lipid interactions in membranes determine cell function and fate, but much remains to be learned about the exact nature of these processes. Membrane-modifying enzymes, cell-surface receptors and transporters are all important targets in pharmaceutical therapy. To better understand lipid function and dynamics, Gerrit van Meer and Kai Simons initiated the EuroMEMBRANE program (part of EuroCORES, funded by the European Research Foundation). According to Gerrit van Meer, the aim of the program is to learn about ‘membrane lipids, where they are and how they behave’. The final EuroMEMBRANE conference in Basel in 2012 (www.lipidsignaling.org) concluded the program, and covered various aspects of membrane biology. Plenary lectures by Pietro De Camilli on ‘Membrane dynamics and phosphoinositide signaling’ and Robin Irvine on ‘Metabolism and functions of PIPs controlling signaling and membrane identity’ embraced the conference program, and the following mini-reviews illustrate a small subset of the themes presented at the conference. The first mini-review by Raiborg et al. describes the function of the class III phosphoinositide 3–kinase Vps34, and how its lipid product phosphatidylinositol 3–phosphate recruits elements of the endosomal sorting machinery, exploiting lipid-binding domains including PX and FYVE domains. In the second mini-review, Solinger and Spang demonstrate how Caenorhabditis elegans genetics may be exploited to determine the ‘to do list’ of endosomal tethering complexes, and integrate these in endosome maturation processes with V–ATPase, the ESCRT complexes, phosphoinositides, and more. The third mini-review by Koren and Bentires-Alj covers the currently available mouse models that are being used to investigate breast cancer development. In tumorigenesis, palmitoylation of oncogenic proteins such as Ras modulates translocation and activation, but the role of hydrophobic post-translational modification of membrane proteins is less obvious. Blaskovic et al. address this issue in their mini-review ‘What does S-palmitoylation do to membrane proteins?‘. Parmryd and Önfelt discuss the importance of membrane curvature in function, but also describe how membrane topology may affect data acquired by various microscopic techniques. In ‘Molecular dynamics simulations of the interactions of medicinal plant extracts and drugs with lipid bilayer membranes’, Kopeć et al. show how compounds affect localized lipid environments. In their mini-review, Volinsky and Kinnunen show that lipids age and are turned over, and that these modifications affect membrane properties and cell fate. Finally, Loizides-Mangold provides insight into how novel technologies such as MS-based lipidomics may be utilized to unravel the complexity of the lipidome. Matthias P. Wymann is Professor at the Department of Biomedicine at the University of Basel (Switzerland). He introduced the first PI3K inhibitor, wortmannin, and has since contributed to the understanding of inhibitor–PI3K interactions, molecular functions and the physiological importance of PI3K. He has initiated European consortia (EU: ACID) to promote the development of isoform-specific PI3K inhibitors, and to validate PI3Ks as drug targets in chronic inflammation, allergy and oncology. With the European Science Foundation (ESF)-funded consortium Tracking of Phosphoinositide Pools (TraPPs), he aims to develop advanced tools to track and modulate phosphoinositide signaling. He is co-founder of PIQUR Therapeutics AG, which is engaged in the development of drugs targeting PI3K and mTOR pathways. Kai Simons received his MD degree from the University of Helsinki, Finland. He then performed postdoctoral research at Rockefeller University in New York. In 1975, Simons moved to the European Molecular Biology Laboratory (EMBL) in Heidelberg, Germany, where he started the Cell Biology Program, which became the focal point for molecular cell biology in Europe. In 2001, Kai Simons moved to Dresden to build up the new Max Planck Institute for Molecular Cell Biology and Genetics, which today is an internationally recognized center of excellence. His recent research has focused on cell membrane organization and function. He has pioneered the concept of lipid rafts as a membrane-organizing principle.

Glucose regulates transcription in yeast through a network of signaling pathways

Addition of glucose to yeast cells increases their growth rate and results in a massive restructuring of their transcriptional output. We have used microarray analysis in conjunction with conditional mutations to obtain a systems view of the signaling network responsible for glucose-induced transcriptional changes. We found that several well-studied signaling pathways—such as Snf1 and Rgt—are responsible for specialized but limited responses to glucose. However, 90% of the glucose-induced changes can be recapitulated by the activation of protein kinase A (PKA) or by the induction of PKB (Sch9). Blocking signaling through Sch9 does not interfere with the glucose response, whereas blocking signaling through PKA does. We conclude that both Sch9 and PKA regulate a massive, nutrient-responsive transcriptional program promoting growth, but that they do so in response to different nutritional inputs. Moreover, activating PKA completely recapitulates the transcriptional growth program in the absence of any increase in growth or metabolism, demonstrating that activation of the growth program results solely from the cell's perception of its nutritional status.

Reading the Hedgehog morphogen gradient by measuring the ratio of bound to unbound Patched protein.

Morphogens are 'form-generating' substances that spread from localized sites of production and specify distinct cellular outcomes at different concentrations. A cell's perception of morphogen concentration is thought to be determined by the number of active receptors, with inactive receptors making little if any contribution. Patched (Ptc), the receptor for the morphogen Hedgehog (Hh), is active in the absence of ligand and blocks the expression of target genes by inhibiting Smoothened (Smo), an essential transducer of the Hh signal. Hh binding to Ptc abrogates the ability of Ptc to inhibit Smo, thereby unleashing Smo activity and inducing target gene expression. Here, we show that a cell's measure of ambient Hh concentration is not determined solely by the number of active (unliganded) Ptc molecules. Instead, we find that Hh-bound Ptc can titrate the inhibitory action of unbound Ptc. Furthermore, we demonstrate that this effect is sufficient to allow normal reading of the Hh gradient in the presence of a form of Ptc that cannot bind the ligand but retains its ability to inhibit Smo. These results support a model in which the ratio of bound to unbound Ptc molecules determines the cellular response to Hh.

Increased Expression of Matrix Metalloproteinase-1 in Osteocytes Precedes Bone Resorption as Stimulated by Disuse: Evidence for Autoregulation of the Cell's Mechanical Environment?

An in vivo animal model of bone adaptation was used to examine a possible role for matrix metalloproteinase-1 in the local mediation of bone remodeling: to corrode the coupling of osteocytes to the matrix in an attempt to autoregulate the cell's perception of its mechanical environment. Twelve young (12-16 months old) skeletally mature turkeys were separated into groups to be studied for stimulus periods of either 3 or 30 days. In each animal, the left ulna was functionally isolated and subjected to either disuse or 3,000 microstrain at 1 Hz for 10 minutes per day. The right ulna remained intact and served as an intra-animal control. No significant differences in bone area were detected at 3 days; however, ulnae subjected to disuse lost 8 +/- 4% (+/-SD) of bone area by 30 days. Over the same period, ulnae subjected to the mechanical stimulus gained 21 +/- 9% of bone area. With use of in situ reverse transcription-polymerase chain reaction, less than 2% of the osteocytes examined from the intact control ulnae stained positively for matrix metalloproteinase-1 mRNA. An antibody raised against matrix metalloproteinase-1 revealed no positively labeled osteocytes in the intact ulnae. This low percentage of osteocytes expressing matrix metalloproteinase-1 mRNA was similar to that seen in ulnae subjected to the osteogenic mechanical stimuli. In contrast, ulnae subjected to either 3 or 30 days of disuse showed evidence of matrix metalloproteinase-1 mRNA activity in a high percentage of osteocytes (89 +/- 5 and 66 +/- 8%, respectively; each time point significantly different from intact ulnae, as well as from each other, p < 0.05). The percentage of osteocytes labeled with the anti-matrix metalloproteinase-1 antibody was also highly elevated following 3 days of disuse (74 +/- 17%). These data demonstrate that an early response of bone to disuse is the upregulation of matrix metalloproteinase-1 activity in osteocytes. It is proposed that this upregulation of collagenase activity is indicative of the cell's degradation of coupling to the matrix, and it thus reflects the osteocyte's regulation of its own mechanical environment. We believe that such autoregulation of the osteocyte's physical environment will accommodate subtle changes in the bone's functional environment without the need to add or resorb bone tissue.

Increased expression of matrix metalloproteinase-1 in osteocytes precedes bone resorption as stimulated by disuse: evidence for autoregulation of the cell's mechanical environment?

An in vivo animal model of bone adaptation was used to examine a possible role for matrix metalloproteinase-1 in the local mediation of bone remodeling: to corrode the coupling of osteocytes to the matrix in an attempt to autoregulate the cell's perception of its mechanical environment. Twelve young (12-16 months old) skeletally mature turkeys were separated into groups to be studied for stimulus periods of either 3 or 30 days. In each animal, the left ulna was functionally isolated and subjected to either disuse or 3,000 microstrain at 1 Hz for 10 minutes per day. The right ulna remained intact and served as an intra-animal control. No significant differences in bone area were detected at 3 days; however, ulnae subjected to disuse lost 8 +/- 4% (+/-SD) of bone area by 30 days. Over the same period, ulnae subjected to the mechanical stimulus gained 21 +/- 9% of bone area. With use of in situ reverse transcription-polymerase chain reaction, less than 2% of the osteocytes examined from the intact control ulnae stained positively for matrix metalloproteinase-1 mRNA. An antibody raised against matrix metalloproteinase-1 revealed no positively labeled osteocytes in the intact ulnae. This low percentage of osteocytes expressing matrix metalloproteinase-1 mRNA was similar to that seen in ulnae subjected to the osteogenic mechanical stimuli. In contrast, ulnae subjected to either 3 or 30 days of disuse showed evidence of matrix metalloproteinase-1 mRNA activity in a high percentage of osteocytes (89 +/- 5 and 66 +/- 8%, respectively; each time point significantly different from intact ulnae, as well as from each other, p < 0.05). The percentage of osteocytes labeled with the anti-matrix metalloproteinase-1 antibody was also highly elevated following 3 days of disuse (74 +/- 17%). These data demonstrate that an early response of bone to disuse is the upregulation of matrix metalloproteinase-1 activity in osteocytes. It is proposed that this upregulation of collagenase activity is indicative of the cell's degradation of coupling to the matrix, and it thus reflects the osteocyte's regulation of its own mechanical environment. We believe that such autoregulation of the osteocyte's physical environment will accommodate subtle changes in the bone's functional environment without the need to add or resorb bone tissue.

A new organelle related to osmoregulation in ultrarapidly frozenPelvetia embryos

Freeze-fracture electron microscopy of the cortical cytoplasm of unfixed, uncryoprotected, ultrarapidly frozen embryos of the marine brown algaPelvetia fastigiata has demonstrated the presence of numerous 0.5-μm diameter, disc-shaped vesicles lying adjacent and nearly parallel to the plasma membrane. Some vesicles are fused with the plasma membrane through a narrow connection; this however appears to be a reversible attachment rather than an intermediate stage in the incorporation of the vesicle into the plasma membrane. The distribution of these connections in the plane of the membrane is not uniform; they tend to occur in patches. The fraction of vesicles that is fused with the plasma membrane at any one time appears to be related to a cell's perception of a stressful hypotonic imbalance between the internal and external concentrations of osmotically active compounds. Thus, a sudden 5% decrease in osmolarity of the artificial seawater medium just before freezing leads to a 38% increase in connections per unit membrane area, while a 20% decrease in osmolarity leads to a 75% increase in connections per unit area. Based on these findings and the corresponding ion-transport studies of R. Nuccitelli and L.F. Jaffe (1976, Planta131, 315-320), we postulate that the disc-shaped vesicles mediate short-term osmoregulation inPelvetia embryos by reversibly inserting chloride channels into the plasma membrane.