Recruitment of Eph receptors into signaling clusters does not require ephrin contact

Eph receptors and their cell membrane-bound ephrin ligands regulate cell positioning and thereby establish or stabilize patterns of cellular organization. Although it is recognized that ephrin clustering is essential for Eph function, mechanisms that relay information of ephrin density into cell biological responses are poorly understood. We demonstrate by confocal time-lapse and fluorescence resonance energy transfer microscopy that within minutes of binding ephrin-A5-coated beads, EphA3 receptors assemble into large clusters. While remaining positioned around the site of ephrin contact, Eph clusters exceed the size of the interacting ephrin surface severalfold. EphA3 mutants with compromised ephrin-binding capacity, which alone are incapable of cluster formation or phosphorylation, are recruited effectively and become phosphorylated when coexpressed with a functional receptor. Our findings reveal consecutive initiation of ephrin-facilitated Eph clustering and cluster propagation, the latter of which is independent of ephrin contacts and cytosolic Eph signaling functions but involves direct Eph-Eph interactions.     

Eph receptor and ephrin function in breast,gut, and skin epithelia

Epithelial cells are tightly coupled together through specialized intercellular junctions, including adherens junctions, desmosomes, tight junctions, and gap junctions. A growing body of evidence suggests epithelial cells also directly exchange information at cell-cell contacts via the Eph family of receptor tyrosine kinases and their membrane-associated ephrin ligands. Ligand-dependent and -independent signaling via Eph receptors as well as reverse signaling through ephrins impact epithelial tissue homeostasis by organizing stem cell compartments and regulating cell proliferation, migration, adhesion, differentiation, and survival. This review focuses on breast, gut, and skin epithelia as representative examples for how Eph receptors and ephrins modulate diverse epithelial cell responses in a context-dependent manner. Abnormal Eph receptor and ephrin signaling is implicated in a variety of epithelial diseases raising the intriguing possibility that this cell-cell communication pathway can be therapeutically harnessed to normalize epithelial function in pathological settings like cancer or chronic inflammation.     

Characterization of loss-of-function and gain-of-function Eph receptor tyrosine kinase signaling in C. elegans axon targeting and cell migration

To understand how our brains function, it is necessary to know how neurons position themselves and target their axons and dendrites to their correct locations. Several evolutionarily conserved axon guidance molecules have been shown to help navigate axons to their correct target site. The Caenorhabditis elegans Eph receptor tyrosine kinase (RTK), VAB-1, has roles in early neuroblast and epidermal cell movements, but its roles in axon guidance are not well understood. Here, we report that mutations that disrupt the VAB-1 Eph receptor tyrosine kinase cause incompletely penetrant defects in axonal targeting and neuronal cell body positioning. The predominant axonal defect in vab-1 mutant animals was an overextension axon phenotype. Interestingly, constitutively active VAB-1 tyrosine kinase signaling caused a lack of axon outgrowth or an early termination phenotype, opposite to the loss-of-function phenotype. The combination of loss-of-function and gain-of-function analyses suggests that the VAB-1 Eph RTK is required for targeting or limiting axons and neuronal cells to specific regions, perhaps by transducing a repellent or stop cue.     

Collagen recognition and transmembrane signalling by discoidin domain receptors

The discoidin domain receptors, DDR1 and DDR2, are two closely related receptor tyrosine kinases that are activated by triple-helical collagen in a slow and sustained manner. The DDRs have important roles in embryo development and their dysregulation is associated with human diseases, such as fibrosis, arthritis and cancer. The extracellular region of DDRs consists of a collagen-binding discoidin (DS) domain and a DS-like domain. The transmembrane region mediates the ligand-independent dimerisation of DDRs and is connected to the tyrosine kinase domain by an unusually long juxtamembrane domain. The major DDR binding site in fibrillar collagens is a GVMGFO motif (O is hydroxyproline), which is recognised by an amphiphilic trench at the top of the DS domain. How collagen binding leads to DDR activation is not understood. GVMGFO-containing triple-helical peptides activate DDRs with the characteristic slow kinetics, suggesting that the supramolecular structure of collagen is not required. Activation can be blocked allosterically by monoclonal antibodies that bind to the DS-like domain. Thus, collagen most likely causes a conformational change within the DDR dimer, which may lead to the formation of larger DDR clusters. This article is part of a Special Issue entitled: Emerging recognition and activation mechanisms of receptor tyrosine kinases.     

Angiotensin II: Biosynthesis,molecular recognition,and signal transduction

Summary 1. Angiotensin II is a well-known vasopressive octapeptide that is the principal end-product of the renin-angiotensin system. In addition to its tonic effect on vascular smooth muscle cells, it also stimulates aldosterone secretion from the adrenals and promotes sodium reabsorption through renal tubular cells.2. These physiological functions have been appreciated for some time, but as details of the molecular and cell biology of the angiotensin response mechanism become understood, it is increasingly apparent that the hormone has a much broader repertoire. Its functional variability is made possible by (i) different enzymatic routes for its generation, (ii) different receptors distributed in different tissues, (iii) different mechanisms for receptor regulation, and (iv) different signal transduction pathways.3. This insight is the direct consequence of advances in pharmacology that led first to inhibitors of angiotensin converting enzyme and later to angiotensin II receptor antagonists. This review looks at the current status of angiotensin biochemistry and physiology and provides a basis for anticipation of future developments.     

Growth factor receptors in helminth parasites: signalling and host-parasite relationships

Parasitic helminths remain major pathogens of both humans and animals throughout the world. The success of helminth infections depends on the capacity of the parasite to counteract host immune responses but also to exploit host-derived signal molecules for its development. Recent progress has been made in the characterization of growth factor receptors of various nematode and flatworm parasites with the demonstration that transforming growth factor beta (TGF-beta), epidermal growth factor (EGF) and insulin receptor signalling pathways are conserved in helminth parasites and potentially implicated in the host-parasite molecular dialogue and parasite development.     

The physiological and pathophysiological role of adiponectin and adiponectin receptors in the peripheral tissues and CNS

Adiponectin is an abundantly expressed adipokine in adipose tissue and has direct insulin sensitizing activity. A decrease in the circulating levels of adiponectin by interactions between genetic factors and environmental factors causing obesity has been shown to contribute to the development of insulin resistance, type 2 diabetes, metabolic syndrome and atherosclerosis. In addition to its insulin sensitizing actions, adiponectin has central actions in the regulation of energy homeostasis. Adiponectin enhances AMP-activated protein kinase activity in the arcuate hypothalamus via its receptor AdipoR1 to stimulate food intake and decreases energy expenditure. We propose a hypothesis on the physiological role of adiponectin: a starvation gene in the course of evolution by promoting fat storage on facing the loss of adiposity.     

Predicted 3-D structures for mouse I7 and rat I7 olfactory receptors and comparison of predicted odor recognition profiles with experiment

The first step in the perception of an odor is the activation of one or more olfactory receptors (ORs) following binding of the odorant molecule to the OR. In order to initiate the process of determining how the molecular level receptor-odorant interactions are related to odor perception, we used the MembStruk computational method to predict the three-dimensional (3-D) structure of the I7 OR for both mouse and rat. We then used the HierDock ligand docking computational method to predict the binding site and binding energy for the library of 56 odorants to these receptors for which experiment response data are now available. We find that the predicted 3-D structures of the mouse and rat I7 OR lead to predictions of odorant binding that are in good agreement with the experimental results, thus validating the accuracy of both the 3-D structure and the predicted binding site. In particular we predict that heptanal and octanal both bind strongly to both mouse and rat I7 ORs, which conflicts with the older literature but agrees with recent experiments. To provide the basis of additional validations of our 3-D structures, we also report the odorant binding site for a new odorant (8-hydroxy-octanal) with a novel functionality designed to bind strongly to mouse I7. Such validated computational methods should be very useful in predicting the structure and function of many other ORs.     

Collagen binding specificity of the discoidin domain receptors: Binding sites on collagens II and III and molecular determinants for collagen IV recognition by DDR1

The discoidin domain receptors, DDR1 and DDR2 are cell surface receptor tyrosine kinases that are activated by triple-helical collagen. While normal DDR signalling regulates fundamental cellular processes, aberrant DDR signalling is associated with several human diseases. We previously identified GVMGFO (O is hydroxyproline) as a major DDR2 binding site in collagens I-III, and located two additional DDR2 binding sites in collagen II. Here we extend these studies to the homologous DDR1 and the identification of DDR binding sites on collagen III. Using sets of overlapping triple-helical peptides, the Collagen II and Collagen III Toolkits, we located several DDR2 binding sites on both collagens. The interaction of DDR1 with Toolkit peptides was more restricted, with DDR1 mainly binding to peptides containing the GVMGFO motif. Triple-helical peptides containing the GVMGFO motif induced DDR1 transmembrane signalling, and DDR1 binding and receptor activation occurred with the same amino acid requirements as previously defined for DDR2. While both DDRs exhibit the same specificity for binding the GVMGFO motif, which is present only in fibrillar collagens, the two receptors display distinct preferences for certain non-fibrillar collagens, with the basement membrane collagen IV being exclusively recognised by DDR1. Based on our recent crystal structure of a DDR2-collagen complex, we designed mutations to identify the molecular determinants for DDR1 binding to collagen IV. By replacing five amino acids in DDR2 with the corresponding DDR1 residues we were able to create a DDR2 construct that could function as a collagen IV receptor.     

Engineered variants of InlB with an additional leucine‐rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex

The physiological relevance of contacts in crystal lattices often remains elusive. This was also the case for the complex between the invasion protein internalin B (InlB) from Listeria monocytogenes and its host cell receptor, the human receptor tyrosine kinase (RTK) MET. InlB is a MET agonist and induces bacterial host cell invasion. Activation of RTKs generally involves ligand‐induced dimerization of the receptor ectodomain. The two currently available crystal structures of the InlB:MET complex show the same arrangement of InlB and MET in a 1:1 complex, but different dimeric 2:2 assemblies. Only one of these 2:2 assemblies is predicted to be stable by a computational procedure. This assembly is mainly stabilized by a contact between the Cap domain of InlB from one and the Sema domain of MET from another 1:1 complex. Here, we probe the physiological relevance of this interaction. We generated variants of the leucine‐rich repeat (LRR) protein InlB by inserting an additional repeat between the first and the second LRR. This should allow formation of the 1:1 complex but disrupt the potential 2:2 complex involving the Cap‐Sema contact due to steric distortions. A crystal structure of one of the engineered proteins showed that it folded properly. Binding affinity to MET was comparable to that of wild‐type InlB. The InlB variant induced MET phosphorylation and cell scatter like wild‐type InlB. These results suggest that the Cap‐Sema interaction is not physiologically relevant and support the previously proposed assembly, in which a 2:2 InlB:MET complex is built around a ligand dimer.     

Targeted therapy for gastric cancer: Molecular pathways and ongoing investigations

Gastric cancer is currently the second leading cause of worldwide cancer mortality. Ongoing collaborative sequencing efforts have highlighted recurrent somatic genomic aberrations in gastric cancer, however, despite advances in characterizing the genomic landscape, there have been few advances in patient outcomes. Prognosis remains poor with a median overall survival of 12 months for advanced disease. The improved survival with trastuzumab, and more recently ramucirumab, underscore the promise of targeted and biologic therapies and the importance of molecular tumor characterization in gastric cancer. Here we review the most frequent actionable alterations in gastric cancer and highlight ongoing clinical investigations attempting to translate biologic understanding into improved clinical outcomes.     

Spectrins: A structural platform for stabilization and activation of membrane channels,receptors and transporters

This review focuses on structure and functions of spectrin as a major component of the membrane skeleton. Recent advances on spectrin function as an interface for signal transduction mediation and a number of data concerning interaction of spectrin with membrane channels, adhesion molecules, receptors and transporters draw a picture of multifaceted protein. Here, we attempted to show the current depiction of multitask role of spectrin in cell physiology. This article is part of a Special Issue entitled: Reciprocal influences between cell cytoskeleton and membrane channels, receptors and transporters. Guest Editor: Jean Claude Hervé.     

Tyrosine kinase and cooperative TGFbeta signaling in the reproductive organs of Schistosoma mansoni

Drug-induced suppression of female schistosome sexual maturation is an auspicious strategy to combat schistosomiasis since the eggs are the causative agent. The establishment of drug targets requires knowledge about the molecular mechanisms that regulate the development of the female reproductive organs, which include vitellarium and ovary. This review summarizes recent studies suggesting tyrosine kinases as important factors for the regulation of female gonad development. In this context, especially cytoplasmatic tyrosine kinases of the Src class seem to play dominant roles. Moreover, experimental data and theoretical concepts are provided supporting a crosstalk between tyrosine kinase and TGFbeta signaling in the production of vitellocytes. Finally, we take advantage from the schistosome genome project to propose a model for the regulation of vitelline-cell production and differentiation.     

Mesdc2 plays a key role in cell-surface expression of Lrp4 and postsynaptic specialization in myotubes

Low-density lipoprotein receptor-related protein 4 (Lrp4) is essential for pre- and post-synaptic specialization at the neuromuscular junction (NMJ), an indispensable synapse between a motor nerve and skeletal muscle. Muscle-specific receptor tyrosine kinase MuSK must form a complex with Lrp4 to organize postsynaptic specialization at NMJs. Here, we show that the chaperon Mesdc2 binds to the intracellular form of Lrp4 and promotes its glycosylation and cell-surface expression. Furthermore, knockdown of Mesdc2 suppresses cell-surface expression of Lrp4, activation of MuSK, and postsynaptic specialization in muscle cells. These results suggest that Mesdc2 plays an essential role in NMJ formation by promoting Lrp4 maturation.     

Key roles for the small leucine-rich proteoglycans in renal and pulmonary pathophysiology

Background

Small leucine-rich proteoglycans (SLRPs) are molecules that have signaling roles in a multitude of biological processes. In this respect, SLRPs play key roles in the evolution of a variety of diseases throughout the human body.

Scope of Review

We will critically review current developments in the roles of SLRPs in several types of disease of the kidney and lungs. Particular emphasis will be given to the roles of decorin and biglycan, the best characterized members of the SLRP gene family.

Major Conclusions

In both renal and pulmonary disorders, SLRPs are essential elements that regulate several pathophysiological processes including fibrosis, inflammation and tumor progression. Decorin has remarkable antifibrotic and antitumorigenic properties and is considered a valuable potential treatment of these diseases. Biglycan can modulate inflammatory processes in lung and renal inflammation and is a potential target in the treatment of inflammatory conditions.

General Significance

SLRPs can serve as either treatment targets or as potential treatment in renal or lung disease. This article is part of a Special Issue entitled Matrix-mediated cell behaviour and properties.     

Cellular functions of NSF: not just SNAPs and SNAREs

N-ethylmaleimide sensitive factor (NSF) is an ATPases associated with various cellular activities protein (AAA), broadly required for intracellular membrane fusion. NSF functions as a SNAP receptor (SNARE) chaperone which binds, through soluble NSF attachment proteins (SNAPs), to SNARE complexes and utilizes the energy of ATP hydrolysis to disassemble them thus facilitating SNARE recycling. While this is a major function of NSF, it does seem to interact with other proteins, such as the AMPA receptor subunit, GluR2, and beta2-AR and is thought to affect their trafficking patterns. New data suggest that NSF may be regulated by transient post-translational modifications such as phosphorylation and nitrosylation. These new aspects of NSF function as well as its role in SNARE complex dynamics will be discussed.