A role for the perlecan protein core in the activation of the keratinocyte growth factor receptor.
Perlecan, a widespread heparan sulphate (HS) proteoglycan, is directly involved in the storing of angiogenic growth factors, mostly members of the fibroblast growth factor (FGF) gene family. We have previously shown that antisense targeting of the perlecan gene causes a reduced growth and responsiveness to FGF7 [also known as keratinocyte growth factor (KGF)] in human cancer cells, and that the perlecan protein core interacts specifically with FGF7. In the present paper, we have investigated human colon carcinoma cells in which the perlecan gene was disrupted by targeted homologous recombination. After screening over 1000 clones, we obtained two clones heterozygous for the null mutation with no detectable perlecan, indicating that the other allele was non-functioning. The perlecan-deficient cells grew more slowly, did not respond to FGF7 with or without the addition of heparin, and were less tumorigenic than control cells. Paradoxically, the perlecan-deficient cells displayed increased FGF7 surface binding. However, the perlecan protein core was required for functional activation of the KGF receptor and downstream signalling. Because heparin could not substitute for perlecan, the HS chains are not critical for FGF7-mediated signalling in this cell system. These results provide the first genetic evidence that the perlecan protein core is a molecular entity implicated in FGF7 binding and activation of its receptor.
Ghiselli
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G
Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, 1020 Locust Street, Philadelphia, PA 19107, USA.
Eichstetter
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I
Iozzo
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RV
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002455:Cell Division / Q000187:drug effects; D003110:Colonic Neoplasms / Q000378:metabolism*; D016373:DNA, Antisense / Q000494:pharmacology; D005343:Fibrinolytic Agents / Q000494:pharmacology; D016220:Fibroblast Growth Factor 1 / Q000378:metabolism; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors / Q000378:metabolism*; D018390:Gene Targeting; D019812:Heparan Sulfate Proteoglycans / Q000502:physiology*; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009374:Neoplasms, Experimental / Q000378:metabolism / Q000473:pathology*; D011485:Protein Binding; D051497:Receptor, Fibroblast Growth Factor, Type 2; D017468:Receptors, Fibroblast Growth Factor / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism / Q000637:transplantation
D016373:DNA, Antisense; C496453:FGF7 protein, human; C496454:Fgf7 protein, mouse; D005343:Fibrinolytic Agents; D019812:Heparan Sulfate Proteoglycans; D017468:Receptors, Fibroblast Growth Factor; D011994:Recombinant Proteins; D016220:Fibroblast Growth Factor 1; D051523:Fibroblast Growth Factor 7; C071927:perlecan; D005346:Fibroblast Growth Factors; D006493:Heparin; D051497:Receptor, Fibroblast Growth Factor, Type 2; C065104:keratinocyte growth factor receptor