title: Mutations in the heparin binding domain of fibronectin in cooperation with the V region induce decreases in pp125(FAK) levels plus proteoglycan-mediated apoptosis via caspases.
abstract: Intact fibronectin (FN) protects cells from apoptosis. When FN is fragmented, specific domains induce proteinase expression in fibroblasts. However, it is not known whether specific domains of FN can also regulate apoptosis. We exposed fibroblasts to four recombinant FN fragments and then assayed for apoptosis using criteria of cellular shape change, condensed nuclear morphology, and DNA fragmentation. The fragments extended from the RGD-containing repeat III10 to III15; they included (V(+)) or excluded (V(-)) the alternatively spliced V region and contained either a mutated (H(-)) or an unmutated (H(+)) heparin binding domain. Only the V(+)H(-) fragment triggered decreases in pp125(FAK) levels and apoptosis, which was rescued by intact FN and inhibitors of caspase-1 and caspase-3. This apoptotic mechanism was mediated by a chondroitin sulfate proteoglycan, since treating cells with chondroitin sulfate or chondroitinase reversed the apoptotic cell shape changes. The alpha4 integrin receptor may also be involved, since using a blocking antibody to alpha4 alone induced apoptotic cell shape changes, whereas co-treatment with this antibody plus V(+)H(+) reversed these effects. These results demonstrate that the V and heparin binding domains of FN modulate pp125(FAK) levels and regulate apoptosis through a chondroitin sulfate proteoglycan- and possibly alpha4 integrin-mediated pathway, which triggers a caspase cascade.
authors.affiliation: Department of Stomatology, University of California, San Francisco, California 94143-0512, USA. ykapila@itsa.ucsf.edu
authors.forename: Y L
authors.initials: YL
authors.lastname: Kapila
authors.forename: S
authors.initials: S
authors.lastname: Wang
authors.forename: P W
authors.initials: PW
authors.lastname: Johnson
mesh_terms: D017398:Alternative Splicing; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001665:Binding Sites; D020170:Caspase 1 / Q000378:metabolism*; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002467:Cell Nucleus / Q000187:drug effects / Q000648:ultrastructure; D002478:Cells, Cultured; D002809:Chondroitin Sulfates / Q000494:pharmacology*; D002810:Chondroitinases and Chondroitin Lyases / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D005353:Fibronectins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006493:Heparin / Q000378:metabolism*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000494:pharmacology*; D010513:Periodontal Ligament / Q000166:cytology / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D061945:Caspase Inhibitors; D015815:Cell Adhesion Molecules; D015853:Cysteine Proteinase Inhibitors; D005353:Fibronectins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D006493:Heparin; D002809:Chondroitin Sulfates; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D020170:Caspase 1; D002810:Chondroitinases and Chondroitin Lyases