title: HSulf-2, an extracellular endoglucosamine-6-sulfatase, selectively mobilizes heparin-bound growth factors and chemokines: effects on VEGF, FGF-1, and SDF-1.
abstract: BACKGROUND
  Heparin/heparan sulfate (HS) proteoglycans are found in the extracellular matrix (ECM) and on the cell surface. A considerable body of evidence has established that heparin and heparan sulfate proteoglycans (HSPGs) interact with numerous protein ligands including fibroblast growth factors, vascular endothelial growth factor (VEGF), cytokines, and chemokines. These interactions are highly dependent upon the pattern of sulfation modifications within the glycosaminoglycan chains. We previously cloned a cDNA encoding a novel human endosulfatase, HSulf-2, which removes 6-O-sulfate groups on glucosamine from subregions of intact heparin. Here, we have employed both recombinant HSulf-2 and the native enzyme from conditioned medium of the MCF-7-breast carcinoma cell line. To determine whether HSulf-2 modulates the interactions between heparin-binding factors and heparin, we developed an ELISA, in which soluble factors were allowed to bind to immobilized heparin.
  
  
  RESULTS
  Our results show that the binding of VEGF, FGF-1, and certain chemokines (SDF-1 and SLC) to immobilized heparin was abolished or greatly diminished by pre-treating the heparin with HSulf-2. Furthermore, HSulf-2 released these soluble proteins from their association with heparin. Native Sulf-2 from MCF-7 cells reproduced all of these activities.
  
  
  CONCLUSIONS
  Our results validate Sulf-2 as a new tool for deciphering the sulfation requirements in the interaction of protein ligands with heparin/HSPGs and expand the range of potential biological activities of this enzyme.
authors.affiliation: Department of Anatomy, Comprehensive Cancer Center, University of California, San Francisco, CA 94143-0452, USA. ihcu@itsa.ucsf.edu
authors.forename: Kenji
authors.initials: K
authors.lastname: Uchimura
authors.forename: Megumi
authors.initials: M
authors.lastname: Morimoto-Tomita
authors.forename: Annette
authors.initials: A
authors.lastname: Bistrup
authors.forename: Jessica
authors.initials: J
authors.lastname: Li
authors.forename: Malcolm
authors.initials: M
authors.lastname: Lyon
authors.forename: John
authors.initials: J
authors.lastname: Gallagher
authors.forename: Zena
authors.initials: Z
authors.lastname: Werb
authors.forename: Steven D
authors.initials: SD
authors.lastname: Rosen
mesh_terms: D000230:Adenocarcinoma / Q000201:enzymology / Q000473:pathology; D001943:Breast Neoplasms / Q000201:enzymology / Q000473:pathology; D045744:Cell Line, Tumor / Q000201:enzymology; D054421:Chemokine CCL21; D054377:Chemokine CXCL12; D019742:Chemokines, CC / Q000378:metabolism; D019743:Chemokines, CXC / Q000378:metabolism*; D017077:Culture Media, Conditioned / Q000737:chemistry; D018076:DNA, Complementary / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D016220:Fibroblast Growth Factor 1 / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D006493:Heparin / Q000378:metabolism*; D006497:Heparan Sulfate / Q000378:metabolism; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism; D009363:Neoplasm Proteins / Q000302:isolation & purification / Q000502:physiology; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000502:physiology; D012710:Serum Albumin, Bovine / Q000378:metabolism*; D013429:Sulfatases; D015238:Sulfotransferases / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
chemical_list: D054421:Chemokine CCL21; D054377:Chemokine CXCL12; D019742:Chemokines, CC; D019743:Chemokines, CXC; D017077:Culture Media, Conditioned; D018076:DNA, Complementary; D016220:Fibroblast Growth Factor 1; D016222:Fibroblast Growth Factor 2; D006493:Heparin; D006497:Heparan Sulfate; D016209:Interleukin-8; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; D012710:Serum Albumin, Bovine; D013429:Sulfatases; D015238:Sulfotransferases; D042461:Vascular Endothelial Growth Factor A; C515478:CCL21 protein, human; C515255:CXCL12 protein, human; C467484:VEGFA protein, human; C470792:SULF2 protein, human