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Human erythroleukemia cells adhere to fibronectin: evidence for a Mr 190,000-receptor protein
Abstract:
Human erythroleukemia cells (K562) adhered rapidly on fibronectin (Fn)-coated growth substratum under serum-free conditions. The adhesion could be quantitatively inhibited by the synthetic peptide Arg-Gly-Asp-Ser (RGDS) and upon hemin-induced differentiation or trypsinization of the cells. Many of the cells also displayed rapid spreading that led to a redistribution of F-actin into spreading edges and in many cells also to a formation of typical actin fibers attaching to the ventral aspect of the cells. The spreading of the cells was inhibited by cytochalasin B but not by microtubule-disrupting drugs, suggesting an active role for the microfilament system in the spreading process. Direct overlay assay of electrophoretically separated polypeptides with 125I-Fn showed that in K562 cells there is a major Mr 190,000 Fn-binding protein that is lost upon differentiation. A similar overlay assay with purified plasma and cellular Fns followed by immunostaining with anti-Fn antibodies revealed a reaction with a similar polypeptide. The binding of Fns on the nitrocellulose sheets could be inhibited and the bound Fn eluted by using the RGDS peptide. From octylglucoside extracts of radioactively surface-labeled cells, distinct Mr 190,000/185,000 membrane glycoproteins bound to Fn-heptapeptide-Sepharose, further suggesting that the Mr 190,000 polypeptide would be the Fn-receptor of the K562 cells.
Insights
Human erythroleukemia cells (K562) rapidly adhere to fibronectin via a specific receptor. This adhesion and cell spreading are mediated by the actin cytoskeleton and are inhibited by the RGDS peptide.
Area of Science:
- Cell biology
- Biochemistry
- Molecular biology
Background:
- Fibronectin (Fn) is a crucial extracellular matrix protein involved in cell adhesion, migration, and differentiation.
- K562 erythroleukemia cells provide a model system to study cell adhesion mechanisms.
Purpose of the Study:
- To identify and characterize the fibronectin receptor on K562 cells.
- To investigate the role of the cytoskeleton in fibronectin-mediated cell adhesion and spreading.
Main Methods:
- Cell adhesion assays using fibronectin-coated substrata.
- Inhibition studies with RGDS peptide, cytochalasin B, and microtubule-disrupting drugs.
- Overlay assays with radiolabeled fibronectin and cell extracts.
- Affinity chromatography using Fn-heptapeptide-Sepharose.
Main Results:
- K562 cells exhibit rapid adhesion and spreading on fibronectin, dependent on the microfilament system.
- A 190,000 Mr fibronectin-binding protein was identified in K562 cells, which is downregulated upon hemin-induced differentiation.
- The RGDS peptide inhibited fibronectin binding, and a Mr 190,000/185,000 membrane glycoprotein was isolated that binds to fibronectin.
Conclusions:
- A Mr 190,000 polypeptide functions as the primary fibronectin receptor in K562 cells.
- Cell spreading involves active participation of the actin cytoskeleton.
- Fibronectin receptor expression is regulated during erythroid differentiation.