Prof. E. Yvonne Jones
The beta propeller architecture of a semaphorin cell guidance cue.
Many major questions in receptor biology centre on how interactions occurring outside the cell trigger signalling inside the cell. To answer such questions for the biomedically relevant signalling systems used in human cells we need to be able to express, purify and structurally analyse membrane spanning proteins. This has been a great challenge for the international structural biology community, landmark structures have been determined for prokaryotic membrane proteins but eukaryotic membrane proteins pose numerous additional problems. The necessary methodologies have only very recently been developed, and these have now started to bear fruit, so this is a very exciting time for the field of receptor research. We are one of relatively few structural biology laboratories worldwide with the necessary expertise in eukaryotic expression systems and along with the groups of my colleagues, Drs Radu Aricescu and Christian Siebold, my group has opportunities for doctoral students to undertake research into the detailed molecular mechanisms of cell surface receptor signalling systems. Current work underway in my laboratory includes studies on cell guidance and adhesion receptors, as well as some collaborations on structural immunology (with Strubi PI Dr Maria Harkiolaki and colleagues at the Weatherall Institute for Molecular Medicine). I would expect to tailor individual doctoral projects according to a research student’s particular interests. A flavour of our work on the extracellular side of the cell surface can be gained from recent publications, current work in the laboratory is now heading into the membrane and increasing involves a spectrum of techniques which seamlessly span from the atomic to the cellular scale.
The core technique in my laboratory is x-ray crystallography, but since our aim is to generate insights which seamlessly span from the detailed atomic structure to the cellular context doctoral students necessarily gain experience of a broad range of methodologies. These include molecular biology, prokaryotic and eukaryotic expression systems (including tissue culture of insect and mammalian cells), protein purification, biophysical techniques (including surface plasmon resonance and analytical ultracentrifugation), crystallization, sychrotron data collection, in silico structural analysis, confocal and electron microscopy. In addition we have extensive networks of interdisciplinary collaborations within Oxford as well as internationally so that research students can interface their results with the broader biomedical context (including when appropriate translational research). Doctoral students have joined the group after undergraduate studies in subjects ranging from clinical medicine to biochemistry, chemistry and physics. All share a passion for adding to our understanding of the molecular mechanisms which underpin biomedical research.
Protein Science & Structural Biology
Project reference number: 114
| Name | Department | Institution | Country | |
|---|---|---|---|---|
| Prof E. Yvonne Jones | Structural Biology | Oxford University | UK | jones-pa@strubi.ox.ac.uk |
2010. An extracellular steric seeding mechanism for Eph-ephrin signaling platform assembly. Nat. Struct. Mol. Biol., 17 (4), pp. 398-402. Read abstract | Read more
Erythropoetin-producing hepatoma (Eph) receptors are cell-surface protein tyrosine kinases mediating cell-cell communication. Upon activation, they form signaling clusters. We report crystal structures of the full ectodomain of human EphA2 (eEphA2) both alone and in complex with the receptor-binding domain of the ligand ephrinA5 (ephrinA5 RBD). Unliganded eEphA2 forms linear arrays of staggered parallel receptors involving two patches of residues conserved across A-class Ephs. eEphA2-ephrinA5 RBD forms a more elaborate assembly, whose interfaces include the same conserved regions on eEphA2, but rearranged to accommodate ephrinA5 RBD. Cell-surface expression of mutant EphA2s showed that these interfaces are critical for localization at cell-cell contacts and activation-dependent degradation. Our results suggest a 'nucleation' mechanism whereby a limited number of ligand-receptor interactions 'seed' an arrangement of receptors which can propagate into extended signaling arrays. Hide abstract
2010. Structural basis of semaphorin-plexin signalling. Nature, 467 (7319), pp. 1118-22. Read abstract | Read more
Cell-cell signalling of semaphorin ligands through interaction with plexin receptors is important for the homeostasis and morphogenesis of many tissues and is widely studied for its role in neural connectivity, cancer, cell migration and immune responses. SEMA4D and Sema6A exemplify two diverse vertebrate, membrane-spanning semaphorin classes (4 and 6) that are capable of direct signalling through members of the two largest plexin classes, B and A, respectively. In the absence of any structural information on the plexin ectodomain or its interaction with semaphorins the extracellular specificity and mechanism controlling plexin signalling has remained unresolved. Here we present crystal structures of cognate complexes of the semaphorin-binding regions of plexins B1 and A2 with semaphorin ectodomains (human PLXNB1(1-2)-SEMA4D(ecto) and murine PlxnA2(1-4)-Sema6A(ecto)), plus unliganded structures of PlxnA2(1-4) and Sema6A(ecto). These structures, together with biophysical and cellular assays of wild-type and mutant proteins, reveal that semaphorin dimers independently bind two plexin molecules and that signalling is critically dependent on the avidity of the resulting bivalent 2:2 complex (monomeric semaphorin binds plexin but fails to trigger signalling). In combination, our data favour a cell-cell signalling mechanism involving semaphorin-stabilized plexin dimerization, possibly followed by clustering, which is consistent with previous functional data. Furthermore, the shared generic architecture of the complexes, formed through conserved contacts of the amino-terminal seven-bladed β-propeller (sema) domains of both semaphorin and plexin, suggests that a common mode of interaction triggers all semaphorin-plexin based signalling, while distinct insertions within or between blades of the sema domains determine binding specificity. Hide abstract
2011. Proteoglycan-specific molecular switch for RPTPσ clustering and neuronal extension. Science, 332 (6028), pp. 484-8. Read abstract | Read more
Heparan and chondroitin sulfate proteoglycans (HSPGs and CSPGs, respectively) regulate numerous cell surface signaling events, with typically opposite effects on cell function. CSPGs inhibit nerve regeneration through receptor protein tyrosine phosphatase sigma (RPTPσ). Here we report that RPTPσ acts bimodally in sensory neuron extension, mediating CSPG inhibition and HSPG growth promotion. Crystallographic analyses of a shared HSPG-CSPG binding site reveal a conformational plasticity that can accommodate diverse glycosaminoglycans with comparable affinities. Heparan sulfate and analogs induced RPTPσ ectodomain oligomerization in solution, which was inhibited by chondroitin sulfate. RPTPσ and HSPGs colocalize in puncta on sensory neurons in culture, whereas CSPGs occupy the extracellular matrix. These results lead to a model where proteoglycans can exert opposing effects on neuronal extension by competing to control the oligomerization of a common receptor. Hide abstract
2011. Modular mechanism of Wnt signaling inhibition by Wnt inhibitory factor 1. Nat. Struct. Mol. Biol., 18 (8), pp. 886-93. Read abstract | Read more
Wnt morphogens control embryonic development and homeostasis in adult tissues. In vertebrates the N-terminal WIF domain (WIF-1(WD)) of Wnt inhibitory factor 1 (WIF-1) binds Wnt ligands. Our crystal structure of WIF-1(WD) reveals a previously unidentified binding site for phospholipid; two acyl chains extend deep into the domain, and the head group is exposed to the surface. Biophysical and cellular assays indicate that there is a WIF-1(WD) Wnt-binding surface proximal to the lipid head group but also implicate the five epidermal growth factor (EGF)-like domains (EGFs I-V) in Wnt binding. The six-domain WIF-1 crystal structure shows that EGFs I-V are wrapped back, interfacing with WIF-1(WD) at EGF III. EGFs II-V contain a heparan sulfate proteoglycan (HSPG)-binding site, consistent with conserved positively charged residues on EGF IV. This combination of HSPG- and Wnt-binding properties suggests a modular model for the localization of WIF-1 and for signal inhibition within morphogen gradients. Hide abstract
2011. A dual binding mode for RhoGTPases in plexin signalling. PLoS Biol., 9 (8), pp. e1001134. Read abstract | Read more
Plexins are cell surface receptors for the semaphorin family of cell guidance cues. The cytoplasmic region comprises a Ras GTPase-activating protein (GAP) domain and a RhoGTPase binding domain. Concomitant binding of extracellular semaphorin and intracellular RhoGTPase triggers GAP activity and signal transduction. The mechanism of this intricate regulation remains elusive. We present two crystal structures of the human Plexin-B1 cytoplasmic region in complex with a constitutively active RhoGTPase, Rac1. The structure of truncated Plexin-B1-Rac1 complex provides no mechanism for coupling RhoGTPase and Ras binding sites. On inclusion of the juxtamembrane helix, a trimeric structure of Plexin-B1-Rac1 complexes is stabilised by a second, novel, RhoGTPase binding site adjacent to the Ras site. Site-directed mutagenesis combined with cellular and biophysical assays demonstrate that this new binding site is essential for signalling. Our findings are consistent with a model in which extracellular and intracellular plexin clustering events combine into a single signalling output. Hide abstract
2007. Structure of a tyrosine phosphatase adhesive interaction reveals a spacer-clamp mechanism. Science, 317 (5842), pp. 1217-20. Read abstract | Read more
Cell-cell contacts are fundamental to multicellular organisms and are subject to exquisite levels of control. Human RPTPmu is a type IIB receptor protein tyrosine phosphatase that both forms an adhesive contact itself and is involved in regulating adhesion by dephosphorylating components of cadherin-catenin complexes. Here we describe a 3.1 angstrom crystal structure of the RPTPmu ectodomain that forms a homophilic trans (antiparallel) dimer with an extended and rigid architecture, matching the dimensions of adherens junctions. Cell surface expression of deletion constructs induces intercellular spacings that correlate with the ectodomain length. These data suggest that the RPTPmu ectodomain acts as a distance gauge and plays a key regulatory function, locking the phosphatase to its appropriate functional location. Hide abstract