Autotaxin, a secreted phosphodiesterase with diverse roles in disease: structural and functional studies
05 / 2011 - 05 / 2016
Autotaxin (ATX) is a secreted, multi-domain, phosphodiesterase that is expressed by many cell types and is present in the circulation. While ATX can hydrolyze nucleotides, it primarily functions as a phospholipase D to convert extracellular LPC (lysophospatidylcholine) into the lipid mediator LPA (lysophosphatidic acid). LPA acts on specific G protein-coupled receptors and thereby stimulates the migration, proliferation and survival of many cell types. The ATX-LPA signalling axis plays important roles in health and disease. Thus, ATX is essential for vascular development and is strongly implicated in inflammation, tumor progression and fibrotic disease, as well as neuropathic pain. These make ATX an attractive target for therapy. Small-molecule inhibitors of ATX are currently being developed in our Institute and elsewhere. While much has been learned about LPA signalling over the years, many key questions concerning ATX are still unanswered. How is ATX activity regulated? What determines the substrate specificity of ATX? What is the function of the non-catalytic domains? Does ATX have non-catalytic signalling functions? How is ATX brought into proximity of cell-surface LPA receptors? Recent evidence indicates that ATX binds with high affinity to heparin as well as to integrins on lymphocytes and platelets, but the mechanism and biological consequences are unknown. We recently achieved a breakthrough by determining the crystal structure of ATX. In the high resolution structure (2.0Å) of the fully functional full-length enzyme, we can clearly see the spatial orientation of the N-terminal somatomedin-B (SMB) domains, the middle catalytic domain (PDE), and the C-terminal nuclease-like domain (NUC). While the active site is clearly visible, the binding of the various ATX substrates and inhibitors needs to be elucidated. Excitingly, one of the SMB domains is positioned in close proximity to the putative binding site of LPC, strongly suggesting a role of the SMB domains in regulating ATX activity or/and substrate presentation. We are now in an excellent position to address many of the outstanding questions on ATX. Specifically, we aim to: " Elucidate how ATX recognizes and binds its substrates on a structural basis, and how the SMB domains participate in that mechanism. We will test our hypothesis that the product, LPA, acts as a "substrate-specifying" factor, preventing ATX from hydrolyzing other substrates such as nucleotides. " Establish how, and through which domain(s), ATX interacts with cell-surface and extracellular matrix molecules, notably integrins and heparin/heparan sulphate. In addition, we will examine the possible non-catalytic signalling functions of ATX itself. To realize our objectives, we will perform structural, biophysical and biochemical experiments for designing cell biological experiments to address ATX function. In this multi-disciplinary approach, structural and biochemical analysis will guide functional studies, e.g. by establishing mutants that have specific substrate recognition properties and can be used in cell biology experiments to dissect the roles of the variable ATX functions. In parallel, functional studies will provide new targets for structural analysis, e.g. dictate which heparin- or integrin- complexes are important and should be crystallized.