Production of cell-associated PDGF-AA by a human sarcoma cell line: evidence for a latent autocrine effect

✍ Scribed by Mozhgan Afrakhte; Monica Mistér; Arne Östman; Bengt Westermark; Ylva Paulsson

• Publisher: John Wiley and Sons
• Year: 1996
• Tongue: French
• Weight: 997 KB
• Volume: 68
• Category: Article
• ISSN: 0020-7136
• DOI: 10.1002/(sici)1097-0215(19961211)68:6<802::aid-ijc19>3.0.co;2-1

✦ Synopsis

The alternative splicing of platelet-derived growth factor A-chain is known to result in 2 different protein products. One variant is encoded by transcripts containing the 69 nts representing exon 6 (PDGF--), and one variant is encoded by transcripts in which exon 6 is excluded (PDGF-NQ. Transfection assays have suggested that the long splice variant of the A-chain is mainly associated with membrane-and matrix-associated heparan sulphate proteoglycans, whereas the shorter variant is soluble. We describe a human sarcoma cell line (U-2 197) that expresses a high level of PDGF-A transcripts. Immunoprecipitations revealed cell-associated protein products of mainly 24.28 and 33 kDa and less abundant forms of 4 0 4 5 kDa, while no PDGF was found in the medium. Analysis of extracellular medium in a radioreceptor assay confirmed that PDGF was not secreted by the U-2 I97 cells. The addition to U-2 197 cultures of a carboxy terminal peptide that specifically competes with the binding of the long splice variant of PDGF-AA to extracellular matrix and cell membranes resulted in the release of 3 PDGF-AA-specific dimeric proteins with molecular masses of 33, 37 and 45 kDa. Furthermore, polymerase chain reaction studies discriminating between the long and the short splice variants of the PDGF-A transcripts revealed that U-2 I97 expressed relatively higher amounts of the long splice variant compared with U-343 MGa CI 26, which is known to secrete PDGF-AA. These cell-associated forms of PDGF, released to the medium by adding carboxy terminal peptide, increased the tyrosine kinase activity of the endogenous PDGF a-receptor.