Signalling pathways in two-component phosphorelay systems

Two-component systems are characterized by phosphotransfer reactions involving histidine and aspartate residues in highly conserved signalling domains. Although the basic principles of signal transduction by these systems have been elucidated, several important aspects, such as their integration into more complex cellular regulatory networks and the molecular basis of the specificity of signal transduction, remain unknown.     

Genetic and molecular analysis of light-regulated plant development

Light regulates many physiological and developmental events in plants through the action of multiple sensory pigment systems. Although our understanding of the regulatory photoreceptors, including phytochromes (that principally absorb red and far-red energy) and blue light receptors, has advanced considerably in the recent past, the mechanisms of light signal transduction in higher plants are poorly understood. To unravel the molecular events associated with light-regulated plant development, a large number of photomorphogenic mutants have been isolated in several different plant species, including Arabidopsis, cucumber, tomato, pea, Brassica and Sorghum, which are either impaired in normal perception of light signal (photoreceptor mutants) or are affected in some specific or a sub-set of phenotypic traits (signal transduction mutants). Their physiological and molecular analysis is proving to be valuable in (1) assigning specific function to discrete phytochrome species, (2) elucidation of elements that constitute the transduction pathway downstream of signal perception, and (3) determining how different photosensory systems regulate many diverse responses. The progress made in the analysis of photomorphogenic mutants, as reviewed in this article, clearly indicates that multiple photoreceptors, either of the same or different class, interact through an intricate network of signal transduction pathways to finally determine the light-dependent phenotype of both monocots and dicots.     

Signal transduction pathways in Synechocystis sp. PCC 6803 and biotechnological implications under abiotic stress

Cyanobacteria have developed various response mechanisms in long evolution to sense and adapt to external or internal changes under abiotic stresses. The signal transduction system of a model cyanobacterium Synechocystis sp. PCC 6803 includes mainly two-component signal transduction systems of eukaryotic-type serine/threonine kinases (STKs), on which most have been investigated at present. These two-component systems play a major role in regulating cell activities in cyanobacteria. More and more co-regulation and crosstalk regulations among signal transduction systems had been discovered due to increasing experimental data, and they are of great importance in corresponding to abiotic stresses. However, mechanisms of their functions remain unknown. Nevertheless, the two signal transduction systems function as an integral network for adaption in different abiotic stresses. This review summarizes available knowledge on the signal transduction network in Synechocystis sp. PCC 6803 and biotechnological implications under various stresses, with focuses on the co-regulation and crosstalk regulations among various stress-responding signal transduction systems.     

Auxin transport and gravitational research: perspectives

Gravity is a fundamental factor which affects all living organisms. Plant development is well adapted to gravity by directing roots downward and shoots upwards. For more than a century, plant biologists have been fascinated to describe the molecular mechanisms underlying the gravitropic response of plants. Important progress towards signal perception, transduction, and response has been made, but new tools are beginning to uncover the regulatory networks for gravitropic control. We summarise recent progress in study of gravitropism and discuss strategies to identify the molecular basis of the gravity response in Arabidopsis thaliana. This will put us on a road towards the molecular systems biology of the Arabidopsis gravitropic response.     

Two isoforms of eukaryotic phospholipase C in Paramecium affecting transport and release of GPI-anchored proteins in vivo

Surface proteins anchored by a glycosylphosphatidylinositol (GPI) residue in the cell membrane are widely distributed among eukaryotic cells. The GPI anchor is cleavable by a phospholipase C (PLC) leading to the release of such surface proteins, and this process is postulated to be essential in several systems. For higher eukaryotes, the responsible enzymes have not been characterized in any detail as yet. Here we characterize six PLCs in the ciliated protozoan, Paramecium, which, in terms of catalytic domains and architecture, all show characteristics of PLCs involved in signal transduction in higher eukaryotes. We show that some of these endogenous PLCs can release GPI-anchored surface proteins in vitro: using RNA_i to reduce PLC expression results in the same effects as the application of PLC inhibitors. With two enzymes, PLC2 and PLC6, RNA_i phenotypes show strong defects in release of GPI-anchored surface proteins in vivo. Moreover, these RNA_i lines also show abnormal surface protein distribution, suggesting that GPI cleavage may influence trafficking of anchored proteins. As we find GFP fusion proteins in the cytosol and in the surface protein extracts, these PLCs obviously show unconventional translocation mechanisms. This is the first molecular data on endogenous Paramecium PLCs with the described properties affecting GPI anchors in vitro and in vivo.     

Role of two-component regulatory systems in intracellular survival of Mycobacterium tuberculosis

The typical two-component regulatory systems (TCSs), consisting of response regulator and histidine kinase, play a central role in survival of pathogenic bacteria under stress conditions such as nutrient starvation, hypoxia, and nitrosative stress. A total of 11 complete paired two-component regulatory systems have been found in Mycobacterium tuberculosis, including a few isolated kinase and regulatory genes. Increasing evidence has shown that TCSs are closely associated with multiple physiological process like intracellular persistence, pathogenicity, and metabolism. This review gives the two-component signal transduction systems in M. tuberculosis and their signal transduction roles in adaption to the environment.     

Expression,purification and activities of the entire family of intact membrane sensor kinases from Enterococcus faecalis

Two-component signal transduction systems are the main mechanism by which bacteria sense and respond to their environment, and their membrane-located histidine protein kinases generally constitute the sensory components of these systems. Relatively little is known about their fundamental mechanisms and precise nature of the molecular signals sensed, because of the technical challenges of producing sufficient quantities of these hydrophobic membrane proteins. This study evaluated the heterologous production, purification and activities of the 16 intact membrane sensor kinases of Enterococcus faecalis. Following the cloning of the genes into expression plasmid pTTQ18His, all but one kinase was expressed successfully in Escherichia coli inner membranes. Purification of the hexa-histidine ‘tagged’ recombinant proteins was achieved for 13, and all but one were verified as intact. Thirteen intact kinases possessed autophosphorylation activity with no added signal when assayed in membrane vesicles or as purified proteins. Signal testing of two functionally-characterized kinases, FsrC and VicK, was successful examplifying the potential use of in vitro activity assays of intact proteins for systematic signal identification. Intact FsrC exhibited an approximately 10-fold increase in activity in response to a two-fold molar excess of synthetic GBAP pheromone, whilst glutathione, and possibly redox potential, were identified for the first time as direct modulators of VicK activity in vitro. The impact of DTT on VicK phosphorylation resulted in increased levels of phosphorylated VicR, the downstream response regulator, thereby confirming the potential of this in vitro approach for investigations of modulator effects on the entire signal transduction process of two-component systems.     

Role of cytokines and extracellular matrix in the regulation of haemopoietic stem cells

The understanding of molecular mechanisms regulating the formation, growth and differentiation of haemopoietic stem cells has advanced considerably recently. Particular progress has been made in defining the cytokines, chemokines and extracellular matrix components which retain and maintain primitive haemopoietic cell populations in bone marrow. Furthermore, signal transduction pathways that are critical for haemopoiesis, both in vivo and in vitro, and that are activated by cytokines have also been identified and further characterised. The importance of these processes has, this year, been exemplified by the phenotypes of mice deficient in key signal transduction proteins and the discovery that mutations in the component proteins of some signalling pathways are linked to human diseases. Significant advances in understanding the molecular mechanisms for mobilisation of stem cells from bone marrow have also been made this year; this has potential importance for bone marrow transplantation.     

Gibberellin signal transduction presents …the SPY who O-GlcNAc'd me

Although the molecular mechanisms by which plants respond to gibberellins are largely unknown, several components of the signal transduction pathway have been identified and a broad outline of how these components act in signal transduction is emerging. One component of the pathway, SPINDLY, is believed to be an O-GlcNAc transferase that post-translationally modifies cytosolic and nuclear proteins by the addition of O-linked N-acetylglucosamine. Although analysis of the properties of O-GlcNAc-modified proteins from animals has led to the hypothesis that this modification is regulatory, it has not been linked to specific signal transduction pathways.     

Two-component signal transduction systems and regulation of virulence factors in <Emphasis Type="Italic">Xanthomonas</Emphasis>: a perspective

Two-component signal transduction systems (TCSTSs), consisting of a histidine kinase and a response regulator, play a critical role in regulating virulence gene expression in Gram-negative phytopathogenic bacteria Xanthomonas spp.. To date, 12 TCSTS genes have been identified, accounting for approximately 10% of the TCSTS genes in each genome that have been experimentally identified to be related to pathogenesis. These TCSTSs modulate the expression of a number of virulence factors through diverse molecular mechanisms such as interacting with DNA, protein-binding and involvement in second messenger metabolism, which generates a high level of regulatory versatility. Here we summarize the current knowledge in this field and discuss the emerging themes and remaining questions that are important in deciphering the signaling network of TCSTSs in Xanthomonas.     

Directed evolution reveals the mechanism of HitRS signaling transduction in Bacillus anthracis

Two component systems (TCSs) are a primary mechanism of signal sensing and response in bacteria. Systematic characterization of an entire TCS could provide a mechanistic understanding of these important signal transduction systems. Here, genetic selections were employed to dissect the molecular basis of signal transduction by the HitRS system that detects cell envelope stress in the pathogen Bacillus anthracis. Numerous point mutations were isolated within HitRS, 17 of which were in a 50-residue HAMP domain. Mutational analysis revealed the importance of hydrophobic interactions within the HAMP domain and highlighted its essentiality in TCS signaling. In addition, these data defined residues critical for activities intrinsic to HitRS, uncovered specific interactions among individual domains and between the two signaling proteins, and revealed that phosphotransfer is the rate-limiting step for signal transduction. Furthermore, this study establishes the use of unbiased genetic selections to study TCS signaling and provides a comprehensive mechanistic understanding of an entire TCS.     

Unravelling the response of poplar (Populus nigra) roots to mechanical stress imposed by bending

Abstract

Mechanical stress is a widespread environmental condition that can be caused by several factors (i.e. gravity, touch, wind, soil density, soil compaction and grazing, slope) and that can severely affect plant stability. In response to mechanical stress and to improve their anchorage, plants have developed complex mechanisms to detect mechanical perturbation and to induce a suite of modifications at anatomical, physiological, biochemical, biophysical and molecular level. Although it is well recognized that one of the primary functions of root systems is to anchor the plant to the soil, root response to mechanical stresses have been investigated mainly at morphological and biomechanical level, whereas investigations about the molecular mechanisms underlying these important alterations are still in an initial stage. We have used an experimental system in which the taproot poplar seedlings are bent to simulate mechanical perturbation to begin investigate the mechanisms involved in root response to mechanical stress. The results reported herein show that, in response to bending, the poplar root changes its morphology by emitting new lateral roots, and its biomechanical properties by increasing the root biomass and lignin synthesis. In addition, using a proteomic approach, we found that several proteins involved in the signal transduction pathway, detoxification and metabolism are up-regulated and/or down-regulated in the bent root. These results provide new insight into the obscure field of woody root response to mechanical stress, and can serve as a basis for future investigations aimed at unravelling the complex mechanism involved in the reaction of root biology to environmental stress.