Structurally encoded intraclass differences in EphA clusters drive distinct cell responses

Functional outcomes of ephrin binding to Eph receptors (Ephs) range from cell repulsion to adhesion. Here we used cell collapse and stripe assays, showing contrasting effects of human ephrinA5 binding to EphA2 and EphA4. Despite equivalent ligand binding affinities, EphA4 triggered greater cell collapse, whereas EphA2-expressing cells adhered better to ephrinA5-coated surfaces. Chimeric receptors showed that the ectodomain is a major determinant of cell response. We report crystal structures of EphA4 ectodomain alone and in complexes with ephrinB3 and ephrinA5. These revealed closed clusters with a dimeric or circular arrangement in the crystal lattice, contrasting with extended arrays previously observed for EphA2 ectodomain. Localization microscopy showed that ligand-stimulated EphA4 induces smaller clusters than does EphA2. Mutant Ephs link these characteristics to interactions observed in the crystal lattices, suggesting a mechanism by which distinctive ectodomain surfaces determine clustering, and thereby signaling, properties. © 2013 Nature America, Inc. All rights reserved

Structural studies of cell surface signalling molecules for neuronal guidance and connectivity

Signal transduction is critical during the lifetime of a neuron as it navigates to reach its targets, forms functional synaptic connections and adjusts the molecular architecture of these connections in an activity-dependent manner. Understanding the molecular organisation of components required for neuronal signalling will provide novel biological insight and can contribute to the design of therapeutics for neurodevelopmental and neurodegenerative disorders. The focus of the thesis is on determining mechanistic molecular details of a number of distinct cell surface systems implicated in neuronal signalling. Crystallographic studies on the cell surface complex between Eph receptor A4 and ephrinA5 contributed to understanding how the modes of higher order arrangements of receptors involved in guidance affect signal transduction across the membrane. A set of structural and biophysical studies addressed the proteoglycan regulation of RPTPσ-TrkCtrans-synaptic interaction and contributed to deciphering the principles of the switch from axonal growth to synapse establishment and formation. A crystallographic and biochemical analysis of the neuronal C1q-like family, enabled mapping their interactions with potential synaptic partners, and guided functional studies aimed at elucidating their roles in the maintenance of synaptic integrity. Preliminary work on the neuronal Sigma-1 receptor chaperone laid the foundations for the structural determination of this receptor.</p

Structural studies of cell surface signalling molecules for neuronal guidance and connectivity

Signal transduction is critical during the lifetime of a neuron as it navigates to reach its targets, forms functional synaptic connections and adjusts the molecular architecture of these connections in an activity-dependent manner. Understanding the molecular organisation of components required for neuronal signalling will provide novel biological insight and can contribute to the design of therapeutics for neurodevelopmental and neurodegenerative disorders. The focus of the thesis is on determining mechanistic molecular details of a number of distinct cell surface systems implicated in neuronal signalling. Crystallographic studies on the cell surface complex between Eph receptor A4 and ephrinA5 contributed to understanding how the modes of higher order arrangements of receptors involved in guidance affect signal transduction across the membrane. A set of structural and biophysical studies addressed the proteoglycan regulation of RPTP&sigma;-TrkCtrans-synaptic interaction and contributed to deciphering the principles of the switch from axonal growth to synapse establishment and formation. A crystallographic and biochemical analysis of the neuronal C1q-like family, enabled mapping their interactions with potential synaptic partners, and guided functional studies aimed at elucidating their roles in the maintenance of synaptic integrity. Preliminary work on the neuronal Sigma-1 receptor chaperone laid the foundations for the structural determination of this receptor.This thesis is not currently available in ORA

Validating business requirements using MAS analysis models

Summarization: This paper presents a method aimed to assist an engineer in transforming agent roles models to a process model, compliant with the XML Process Definition Language (XPDL) portable standard. This method bridges the gap between software engineers and the business world by allowing a Multi-Agent System (MAS) analysis model, such as the Gaia or ASEME Methodologies role models, to be represented as a business process model. Thus, on one hand, the software engineer can employ available tools to validate specific properties of the modeled system even before its final implementation, and, on the other hand, a business partner has greater potential to comprehend the system being modeled. The method includes a tool for aiding the engineer in the transformation process. This tool uses a recursive algorithm for automating the transformation process and guides the user to dynamically integrate two or more agent roles in a process model with multiple pools. The tool usage is demonstrated through a running example, based on a real world project. Simulations of the defined agent roles can be used to a) validate the system requirements and b) determine how it could scale. This way, developers and managers can configure processes’ parameters and identify and resolve risks early in their project.Παρουσιάστηκε στο: 14th International Conference on Autonomous Agents and Multi-Agent System

Validating requirements using gaia roles models

Summarization: This paper presents a method aimed to assist an engineer in transforming agent roles models to a process model. Thus, the software engineer can employ available tools to validate specific properties of the modeled system even before its final implementation. The method includes a tool for aiding the engineer in the transformation process. This tool uses a recursive algorithm for automating the transformation process and guides the user to dynamically integrate two or more agent roles in a process model with multiple pools. The tool usage is demonstrated through a running example, based on a real world project. Simulations of the defined agent roles can be used to a) validate the system requirements and b) determine how it could scale. This way, developers and managers can configure processes’ parameters and identify and resolve risks early in their project.Παρουσιάστηκε στο: Workshop on Engineering Multi-Agent System

Targeting phosphatase-dependent proteoglycan switch for rheumatoid arthritis therapy.

Despite the availability of several therapies for rheumatoid arthritis (RA) that target the immune system, a large number of RA patients fail to achieve remission. Joint-lining cells, called fibroblast-like synoviocytes (FLS), become activated during RA and mediate joint inflammation and destruction of cartilage and bone. We identify RPTPσ, a transmembrane tyrosine phosphatase, as a therapeutic target for FLS-directed therapy. RPTPσ is reciprocally regulated by interactions with chondroitin sulfate or heparan sulfate containing extracellular proteoglycans in a mechanism called the proteoglycan switch. We show that the proteoglycan switch regulates FLS function. Incubation of FLS with a proteoglycan-binding RPTPσ decoy protein inhibited cell invasiveness and attachment to cartilage by disrupting a constitutive interaction between RPTPσ and the heparan sulfate proteoglycan syndecan-4. RPTPσ mediated the effect of proteoglycans on FLS signaling by regulating the phosphorylation and cytoskeletal localization of ezrin. Furthermore, administration of the RPTPσ decoy protein ameliorated in vivo human FLS invasiveness and arthritis severity in the K/BxN serum transfer model of RA. Our data demonstrate that FLS are regulated by an RPTPσ-dependent proteoglycan switch in vivo, which can be targeted for RA therapy. We envision that therapies targeting the proteoglycan switch or its intracellular pathway in FLS could be effective as a monotherapy or in combination with currently available immune-targeted agents to improve control of disease activity in RA patients

Targeting phosphatase-dependent proteoglycan switch for rheumatoid arthritis therapy

Despite the availability of several therapies for rheumatoid arthritis (RA) that target the immune system, a large number of RA patients fail to achieve remission. Joint-lining cells, called fibroblast-like synoviocytes (FLS), become activated during RA and mediate joint inflammation and destruction of cartilage and bone. We identify RPTPσ, a transmembrane tyrosine phosphatase, as a therapeutic target for FLS-directed therapy. RPTPσ is reciprocally regulated by interactions with chondroitin sulfate or heparan sulfate containing extracellular proteoglycans in a mechanism called the proteoglycan switch. We show that the proteoglycan switch regulates FLS function. Incubation of FLS with a proteoglycan-binding RPTPσ decoy protein inhibited cell invasiveness and attachment to cartilage by disrupting a constitutive interaction between RPTPσ and the heparan sulfate proteoglycan syndecan-4. RPTPσ mediated the effect of proteoglycans on FLS signaling by regulating the phosphorylation and cytoskeletal localization of ezrin. Furthermore, administration of the RPTPσ decoy protein ameliorated in vivo human FLS invasiveness and arthritis severity in the K/BxN serum transfer model of RA. Our data demonstrate that FLS are regulated by an RPTPσ-dependent proteoglycan switch in vivo, which can be targeted for RA therapy. We envision that therapies targeting the proteoglycan switch or its intracellular pathway in FLS could be effective as a monotherapy or in combination with currently available immune-targeted agents to improve control of disease activity in RA patients