Generic signal-specific responses: cytokinin and context-dependent cellular responses

The phytohormone cytokinin triggers numerous and diverse responses during the plant life cycle via a two-component phosphorelay signalling system. Each step of the signalling cascade is supported by a gene family comprising several members. While functional redundancy is observed among family members, additional gene-specific functions encoded by cis-regulatory and coding sequence of individual family members have been described and contribute to specificity in signalling output. In addition, the cellular context of the signal-receiving cell affects the response triggered. Recent studies in Arabidopsis have demonstrated how expression of cytokinin signalling components predefines a spatiotemporal map of signalling sensitivity, which causes local signal amplification and attenuation. In summary, the specific interpretation of cytokinin signalling is affected by an orchestrated interplay of signalling genes and cellular context.     

G protein-coupled receptor signalling and cross-talk: achieving rapidity and specificity

Activation of a given type of G protein-coupled receptor (GPCR) triggers a limited set of signalling events in a very rapid and specific manner. The classical paradigm of GPCR signalling was rather linear and sequential. Emerging evidence, however, has revealed that this is only a part of the complex signalling mediated by GPCR. Propagation of GPCR signalling involves cross-regulation of many but specific pathways, including cross-talks between different GPCRs as well as with other signalling pathways. Moreover, it is increasingly apparent that GPCRs can activate both heterotrimeric G protein-dependent and G protein-independent signalling pathways. In this review, we discuss how the signallings initiated by GPCRs achieve rapidity as well as specificity, and how the GPCRs can cross-regulate other specific signalling pathways at the same time. New concepts regarding GPCR signalling have been arising to address this issue, which include multiprotein signalling complex and signalling compartment in microdomain concepts that enable close colocalization or even contact among the proteins engaged in the specific signal transduction. The final outcome of a stimulation of GPCR will thus be the sum of its own specific set of intracellular signalling pathways it regulates.     

Mitotic and mitogenic Wnt signalling

Canonical Wnt signalling plays an important role in development, tissue homeostasis, and cancer. At the cellular level, canonical Wnt signalling acts by regulating cell fate, cell growth, and cell proliferation. With regard to proliferation, there is increasing evidence for a complex interaction between canonical Wnt signalling and the cell cycle. Mitogenic Wnt signalling regulates cell proliferation by promoting G1 phase. In mitosis, components of the Wnt signalling cascade function directly in spindle formation. Moreover, Wnt signalling is strongly activated in mitosis, suggesting that 'mitotic Wnt signalling' plays an important role to orchestrate a cell division program. Here, we review the complex interplay between Wnt signalling and the cell cycle.     

Wingless modulates the effects of dominant negative notch molecules in the developing wing of Drosophila

The development and patterning of the wing in Drosophila relies on a sequence of cell interactions molecularly driven by a number of ligands and receptors. Genetic analysis indicates that a receptor encoded by the Notch gene and a signal encoded by the wingless gene play a number of interdependent roles in this process and display very strong functional interactions. At certain times and places, during wing development, the expression of wingless requires Notch activity and that of its ligands Delta and Serrate. This has led to the proposal that all the interactions between Notch and wingless can be understood in terms of this regulatory relationship. Here we have tested this proposal by analysing interactions between Delta- and Serrate-activated Notch signalling and Wingless signalling during wing development and patterning. We find that the cell death caused by expressing dominant negative Notch molecules during wing development cannot be rescued by coexpressing Nintra. This suggests that the dominant negative Notch molecules cannot only disrupt Delta and Serrate signalling but can also disrupt signalling through another pathway. One possibility is the Wingless signalling pathway as the cell death caused by expressing dominant negative Notch molecules can be rescued by activating Wingless signalling. Furthermore, we observe that the outcome of the interactions between Notch and Wingless signalling differs when we activate Wingless signalling by expressing either Wingless itself or an activated form of the Armadillo. For example, the effect of expressing the activated form of Armadillo with a dominant negative Notch on the patterning of sense organ precursors in the wing resembles the effects of expressing Wingless alone. This result suggests that signalling activated by Wingless leads to two effects, a reduction of Notch signalling and an activation of Armadillo.     

Gradients of signalling in the developing limb

The developing limb is one of the first systems where it was proposed that a signalling gradient is involved in pattern formation. This gradient for specifying positional information across the antero-posterior axis is based on Sonic hedgehog signalling from the polarizing region. Recent evidence suggests that Sonic hedgehog signalling also specifies positional information across the antero-posterior axis by a timing mechanism acting in parallel with graded signalling. The progress zone model for specifying proximo-distal pattern, involving timing to provide cells with positional information, continues to be challenged, and there is further evidence that graded signalling by retinoic acid specifies the proximal part of the limb. Other recent papers present the first evidence that gradients of signalling by Wnt5a and FGFs govern cell behaviour involved in outgrowth and morphogenesis of the developing limb.     

Insulin signalling: metabolic pathways and mechanisms for specificity.

Biological actions of insulin are mediated by the insulin receptor, a member of a large family of receptor tyrosine kinases (RTK). Signal transduction by the insulin receptor follows a paradigm for RTK signalling. Many intracellular signalling molecules contain multiple modular domains that mediate protein-protein interactions and participate in the formation of signalling complexes. Phosphorylation cascades are also a prominent feature of RTK signalling. Distal pathways are difficult to dissect because branching paths emerge from downstream effectors and several upstream inputs converge upon single branch points. Thus, insulin action is determined by complicated signalling networks rather than simple linear pathways. Interestingly, many signalling molecules downstream from the insulin receptor are also activated by a plethora of RTKs. Therefore, mechanisms that generate specificity are required. In this review we discuss recent advances in the elucidation of specific metabolic insulin signalling pathways related to glucose transport, one of the most distinctive biological actions of insulin. We also present examples of potential mechanisms underlying specificity in insulin signalling including interactions between multiple branching pathways, subcellular compartmentalization, tissue-specific expression of key effectors and modulation of signal frequency and amplitude.     

Bacterial signal transduction network in a genomic perspective

Bacterial signalling network includes an array of numerous interacting components that monitor environmental and intracellular parameters and effect cellular response to changes in these parameters. The complexity of bacterial signalling systems makes comparative genome analysis a particularly valuable tool for their studies. Comparative studies revealed certain general trends in the organization of diverse signalling systems. These include (i) modular structure of signalling proteins; (ii) common organization of signalling components with the flow of information from N-terminal sensory domains to the C-terminal transmitter or signal output domains (N-to-C flow); (iii) use of common conserved sensory domains by different membrane receptors; (iv) ability of some organisms to respond to one environmental signal by activating several regulatory circuits; (v) abundance of intracellular signalling proteins, typically consisting of a PAS or GAF sensor domains and various output domains; (vi) importance of secondary messengers, cAMP and cyclic diguanylate; and (vii) crosstalk between components of different signalling pathways. Experimental characterization of the novel domains and domain combinations would be needed for achieving a better understanding of the mechanisms of signalling response and the intracellular hierarchy of different signalling pathways.     

The classic signalling and trans‐signalling of interleukin‐6 are both injurious in podocyte under high glucose exposure

Interleukin‐6 (IL‐6) is a multifunctional cytokine that employs IL‐6 classic and trans‐signalling pathways, and these two signal channels execute different or even opposite effects in certain diseases. As a cardinal event of diabetic kidney disease (DKD), whether the podocyte abnormalities are associated with IL‐6 signalling, especially classic or trans‐signalling respectively, remains unclear. In this study, we identified that the circulatory IL‐6, soluble IL‐6R (sIL‐6R) and soluble glycoprotein 130 (sgp130) levels are elevated in patients with DKD. The expressions of membrane‐bound IL‐6R (mIL‐6R), sIL‐6R and gp130 are enhanced in kidney cortex of diabetic mice accompanying with activated STAT3 by tyrosine 705 residue phosphorylation, while not serine 727. Above data infer both classic signalling and trans‐signalling of IL‐6 are activated during DKD. In cultured podocyte, high glucose (HG) up‐regulates the expression of mIL‐6R and gp130, as well as STAT3 tyrosine 705 phosphorylation, in a time‐dependent manner. Entirely blocking IL‐6 signalling by gp130 shRNA, gp130 or IL‐6 neutralizing antibodies attenuates HG‐induced podocyte injury. Interestingly, either inhibiting IL‐6 classic signalling by mIL‐6R shRNA or suppressing its trans‐signalling using sgp130 protein dramatically alleviates HG‐induced podocyte injury, suggesting both IL‐6 classic signalling and trans‐signalling play a detrimental role in HG‐induced podocyte injury. Additionally, activation of IL‐6 classic or trans‐signalling aggravates podocyte damage in vitro. In summary, our observations demonstrate that the activation of either IL‐6 classic or trans‐signalling advances podocyte harming under hyperglycaemia. Thus, suppressing IL‐6 classic and trans‐signalling simultaneously may be more beneficial in podocyte protection and presents a novel therapeutic target for DKD.     

EGF receptor signalling protects smooth-cuticle cells from apoptosis during Drosophila ventral epidermis development

Patterning of the Drosophila ventral epidermis is a tractable model for understanding the role of signalling pathways in development. Interplay between Wingless and EGFR signalling determines the segmentally repeated pattern of alternating denticle belts and smooth cuticle: spitz group genes, which encode factors that stimulate EGFR signalling, induce the denticle fate, while Wingless signalling antagonizes the effect of EGFR signalling, allowing cells to adopt the smooth-cuticle fate. Medial fusion of denticle belts is also a hallmark of spitz group genes, yet its underlying cause is unknown. We have studied this phenotype and discovered a new function for EGFR signalling in epidermal patterning. Smooth-cuticle cells, which are receiving Wingless signalling, are nevertheless dependent on EGFR signalling for survival. Reducing EGFR signalling results in apoptosis of smooth-cuticle cells between stages 12 and 14, bringing adjacent denticle regions together to result in denticle belt fusions by stage 15. Multiple factors stimulate EGFR signalling to promote smooth-cuticle cell survival: in addition to the spitz group genes, Rhomboid-3/roughoid, but not Rhomboid-2 or -4, and the neuregulin-like ligand Vein also function in survival signalling. Pointed mutants display the lowest frequency of fusions, suggesting that EGFR signalling may inhibit apoptosis primarily at the post-translational level. All ventral epidermal cells therefore require some level of EGFR signalling; high levels specify the denticle fate, while lower levels maintain smooth-cuticle cell survival. This strategy might guard against developmental errors, and may be conserved in mammalian epidermal patterning.     

Endoglin promotes endothelial cell proliferation and TGF-beta/ALK1 signal transduction

Endoglin is a transmembrane accessory receptor for transforming growth factor-beta (TGF-beta) that is predominantly expressed on proliferating endothelial cells in culture and on angiogenic blood vessels in vivo. Endoglin, as well as other TGF-beta signalling components, is essential during angiogenesis. Mutations in endoglin and activin receptor-like kinase 1 (ALK1), an endothelial specific TGF-beta type I receptor, have been linked to the vascular disorder, hereditary haemorrhagic telangiectasia. However, the function of endoglin in TGF-beta/ALK signalling has remained unclear. Here we report that endoglin is required for efficient TGF-beta/ALK1 signalling, which indirectly inhibits TGF-beta/ALK5 signalling. Endothelial cells lacking endoglin do not grow because TGF-beta/ALK1 signalling is reduced and TGF-beta/ALK5 signalling is increased. Surviving cells adapt to this imbalance by downregulating ALK5 expression in order to proliferate. The ability of endoglin to promote ALK1 signalling also explains why ectopic endoglin expression in endothelial cells promotes proliferation and blocks TGF-beta-induced growth arrest by indirectly reducing TGF-beta/ALK5 signalling. Our results indicate a pivotal role for endoglin in the balance of ALK1 and ALK5 signalling to regulate endothelial cell proliferation.     

Transient downregulation of Bmp signalling induces extra limbs in vertebrates

Bone morphogenetic protein (Bmp) signalling has been implicated in setting up dorsoventral patterning of the vertebrate limb and in its outgrowth. Here, we present evidence that Bmp signalling or, more precisely, its inhibition also plays a role in limb and fin bud initiation. Temporary inhibition of Bmp signalling either by overexpression of noggin or using a synthetic Bmp inhibitor is sufficient to induce extra limbs in the Xenopus tadpole or exogenous fins in the Danio rerio embryo, respectively. We further show that Bmp signalling acts in parallel with retinoic acid signalling, possibly by inhibiting the known limb-inducing gene wnt2ba.     

Amino acid signalling and the integration of metabolism

It has become clear in recent years that amino acids are not only important as substrates for various metabolic pathways but that they can also activate a nutrient-sensitive, mTOR-mediated, signalling pathway in synergy with insulin. Leucine is the most effective amino acid in this regard. The signalling pathway is antagonised by AMP-activated protein kinase. Amino acid signalling stimulates protein synthesis and inhibits (autophagic) proteolysis. In addition, many amino acids cause an increase in cell volume. Cell swelling per se stimulates synthesis of protein, glycogen, and lipid, in part by further stimulating signalling and in part by unrelated mechanisms. Amino acids also stimulate signalling in beta-cells and stimulate beta-cell growth and proliferation. This results in increased production of insulin, which enhances the anabolic (and anti-catabolic) properties of amino acids. Finally, amino acid-dependent signalling controls the production of leptin by adipocytes, and thus contributes to the regulation of appetite.     

Lysosomal positioning coordinates cellular nutrient responses

mTOR (mammalian target of rapamycin) signalling and macroautophagy (henceforth autophagy) regulate numerous pathological and physiological processes, including cellular responses to altered nutrient levels. However, the mechanisms regulating mTOR and autophagy remain incompletely understood. Lysosomes are dynamic intracellular organelles intimately involved both in the activation of mTOR complex 1 (mTORC1) signalling and in degrading autophagic substrates. Here we report that lysosomal positioning coordinates anabolic and catabolic responses with changes in nutrient availability by orchestrating early plasma-membrane signalling events, mTORC1 signalling and autophagy. Activation of mTORC1 by nutrients correlates with its presence on peripheral lysosomes that are physically close to the upstream signalling modules, whereas starvation causes perinuclear clustering of lysosomes, driven by changes in intracellular pH. Lysosomal positioning regulates mTORC1 signalling, which in turn influences autophagosome formation. Lysosome positioning also influences autophagosome-lysosome fusion rates, and thus controls autophagic flux by acting at both the initiation and termination stages of the process. Our findings provide a physiological role for the dynamic state of lysosomal positioning in cells as a coordinator of mTORC1 signalling with autophagic flux.     

Graded interference with FGF signalling reveals its dorsoventral asymmetry at the mid-hindbrain boundary

Signalling by fibroblast growth factors (FGFs) at the mid-hindbrain boundary (MHB) is of central importance for anteroposterior neural patterning from the isthmic organiser. Graded suppression of FGF signalling by increasing amounts of a dominant negative FGF receptor provides evidence that in addition to anteroposterior patterning, FGF signalling is also involved in patterning along the dorsoventral axis at the MHB. FGF signalling at the MHB is required for the activation of the HH target gene spalt at the MHB. Our results indicate that FGF signalling mediates the competence of the MHB to activate spalt in response to SHH. This interdependence of the two signalling pathways is also found in the outbudding optic vesicle where HH requires functional FGF signalling to activate spalt in the proximal eye region.     

Light signaling mediated by PAS domain-containing proteins in Xanthomonas campestris pv. campestris

Per-ARNT-Sim (PAS) domains are important signalling modules that possibly monitor changes in various stimuli such as light. For the majority of PAS domains that have been identified by sequence similarity, the biological function of the signalling pathways has not yet been experimentally investigated.Thirty-three PAS proteins were discovered in Xanthomonas campestris pv. campestris(Xcc) by genome/proteome analysis. Thirteen PAS proteins were identified as contributing to light signalling and Xcc growth, motility or virulence using molecular genetics and bioinformatics methods. The PAS domains played important roles in light signalling to regulate the growth, motility and virulence of Xcc. They might be regulated by not only light quality (wavelength)but also quantity (intensity) as potential light-signalling components. Evaluating the light wavelength, three light-signalling types of PAS proteins in Xcc were shown to be involved in blue light signalling, tricolour (blue, red and far red)signalling or red/far-red signalling. This showed that Xcc had evolved a complicated light-signalling system to adapt to a complex environment.     

Signal transduction mechanisms of CD137 ligand in human monocytes

Bidirectional signalling, i.e. simultaneous signalling through a receptor as well as its cell surface-bound ligand has been identified for several members of the TNF and TNF receptor family members. Reverse signalling through the ligands offers the advantage of an immediate feed-back and a more precise fine tuning of biological responses. Little is known about the molecular nature of reverse signalling through the ligands. CD137 ligand, member of the TNF family is expressed on monocytes and induces activation, migration, prolongation of survival and proliferation of monocytes. Here we show that reverse signalling by CD137 ligand is mediated by protein tyrosine kinases, p38 mitogen activated protein kinase (MAPK), extracellular signal-regulated kinase (ERK)1,2, MAP/ERK kinase (MEK), Phosphoinositide-3-kinase (PI3-K) and protein kinase A (PKA) but not by protein kinase C (PKC). This study also shows that reverse signalling relies on the same signal transduction molecules as signalling through classical receptors and is in its nature not different from it.     

The role of calcium in ABA-induced gene expression and stomatal movements

There is much interest in the transduction pathways by which abscisic acid (ABA) regulates stomatal movements (ABA-turgor signalling) and by which this phytohormone regulates the pattern of gene expression in plant cells (ABA-nuclear signalling). A number of second messengers have been identified in both the ABA-turgor and ABA-nuclear signalling pathways. A major challenge is to understand the architecture of ABA-signalling pathways and to determine how the ABA signal is coupled to the appropriate response. We have investigated whether separate Ca2+-dependent and -independent ABA-signalling pathways are present in guard cells. Our data suggest that increases in [Ca2+]i are a common component of the guard cell ABA-turgor and ABA-nuclear signalling pathways. The effects of Ca2+ antagonists on ABA-induced stomatal closure and the ABA-responsive CDeT6-19 gene promoter suggest that Ca2+ is involved in both ABA-turgor signalling and ABA-nuclear signalling in guard cells. However, the sensitivity of these pathways to alterations in the external calcium concentration differ, suggesting that the ABA-nuclear and ABA-turgor signalling pathways are not completely convergent. Our data suggest that whilst Ca2+-independent signalling elements are present in the guard cell, they do not form a completely separate Ca2+-independent ABA-signalling pathway.