title: The polybasic insertion in autotaxin α confers specific binding to heparin and cell surface heparan sulfate proteoglycans.
abstract: Autotaxin (ATX) is a secreted lysophospholipase D that generates the lipid mediator lysophosphatidic acid (LPA), playing a key role in diverse physiological and pathological processes. ATX exists in distinct splice variants, but isoform-specific functions remain elusive. Here we characterize the ATXα isoform, which differs from the canonical form (ATXβ) in having a 52-residue polybasic insertion of unknown function in the catalytic domain. We find that the ATXα insertion is susceptible to cleavage by extracellular furin-like endoproteases, but cleaved ATXα remains structurally and functionally intact due to strong interactions within the catalytic domain. Through ELISA and surface plasmon resonance assays, we show that ATXα binds specifically to heparin with high affinity (K(d) ~10(-8) M), whereas ATXβ does not; furthermore, heparin moderately enhanced the lysophospholipase D activity of ATXα. We further show that ATXα, but not ATXβ, binds abundantly to SKOV3 carcinoma cells. ATXα binding was abolished after treating the cells with heparinase III, but not after chondroitinase treatment. Thus, the ATXα insertion constitutes a cleavable heparin-binding domain that mediates interaction with heparan sulfate proteoglycans, thereby targeting LPA production to the plasma membrane.
authors.affiliation: Division of Cell Biology, The Netherlands Cancer Institute, 1066CX Amsterdam, The Netherlands.
authors.forename: Anna J S
authors.initials: AJ
authors.lastname: Houben
authors.forename: Xander M R
authors.initials: XM
authors.lastname: van Wijk
authors.forename: Laurens A
authors.initials: LA
authors.lastname: van Meeteren
authors.forename: Leonie
authors.initials: L
authors.lastname: van Zeijl
authors.forename: Els M A
authors.initials: EM
authors.lastname: van de Westerlo
authors.forename: Jens
authors.initials: J
authors.lastname: Hausmann
authors.forename: Alexander
authors.initials: A
authors.lastname: Fish
authors.forename: Anastassis
authors.initials: A
authors.lastname: Perrakis
authors.forename: Toin H
authors.initials: TH
authors.lastname: van Kuppevelt
authors.forename: Wouter H
authors.initials: WH
authors.lastname: Moolenaar
mesh_terms: D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000378:metabolism; D002465:Cell Movement; D018360:Crystallography, X-Ray / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D057809:HEK293 Cells; D019812:Heparan Sulfate Proteoglycans / Q000737:chemistry*; D006493:Heparin / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D008055:Lipids / Q000737:chemistry; D008246:Lysophospholipids / Q000737:chemistry; D008856:Microscopy, Fluorescence / Q000379:methods; D008969:Molecular Sequence Data; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D015398:Signal Transduction; D000099189:Lysophospholipase D
chemical_list: D019812:Heparan Sulfate Proteoglycans; D006493:Heparin; D008055:Lipids; D008246:Lysophospholipids; D010727:Phosphoric Diester Hydrolases; D000099189:Lysophospholipase D; C032881:lysophosphatidic acid