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Identification and characterization of an intracellular protein complex that binds fibroblast growth factor-2 in
E Chevet1, G Lemaître, K Cailleret
1UER de Science, Université Paris XII, 61 avenue du Général De Gaulle, 94010 Créteil Cédex, France.
Abstract:
The fibroblast growth factor (FGF) family is composed of polypeptides with sequence identity which signal through transmembrane tyrosine kinase receptors. We report here the purification from bovine brain microsomes of an FGF-2-binding complex composed of three proteins of apparent molecular masses 150 kDa, 79 kDa and 46 kDa. Only the 150 kDa and 79 kDa proteins bound FGF-2 in cross-linking and ligand-blotting experiments. Binding of FGF-2 to p79 is enhanced in the presence of calcium. Peptide sequences allowed the identification of p150 and the cloning of the cDNAs encoding p79 and p46. The deduced amino acid sequence of p79 reveals high similarity to those of gastrin-binding protein and mitochondrial enoyl-CoA hydratase/hydroxyacyl-CoA dehydrogenase. p46 is similar to mitochondrial ketoacyl-CoA thiolase. Stable transfection of FR3T3 rat fibroblast cells with p79 cDNA analysed by electron microscopy following immunolabelling of ultra-thin cryosections revealed a localization of p79 in the secretory pathway, mainly in the endoplasmic reticulum and the Golgi region, where it is specifically associated with the molecular chaperone calnexin. In vivo a protein similar to the Golgi protein MG-160 forms a complex with FGF-2 and p79.
Insights
Researchers purified a fibroblast growth factor-2 (FGF-2) binding complex from bovine brain microsomes. This complex contains three proteins, with p150 and p79 directly binding FGF-2, suggesting a novel signaling pathway.
Area of Science:
- Cell biology
- Molecular biology
- Biochemistry
Background:
- The fibroblast growth factor (FGF) family comprises signaling polypeptides that interact with transmembrane tyrosine kinase receptors.
- Understanding FGF-2 interactions is crucial for deciphering cellular signaling pathways.
Purpose of the Study:
- To identify and characterize proteins that bind fibroblast growth factor-2 (FGF-2).
- To elucidate the molecular composition and cellular localization of an FGF-2 binding complex.
Main Methods:
- Purification of FGF-2 binding complex from bovine brain microsomes.
- Cross-linking and ligand-blotting assays to identify FGF-2 binding proteins.
- Peptide sequencing and cDNA cloning for protein identification.
- Stable cell transfection and immunoelectron microscopy for localization studies.
Main Results:
- A three-protein complex (150 kDa, 79 kDa, 46 kDa) binding FGF-2 was purified.
- Proteins p150 and p79 were identified as direct FGF-2 binders; p79 binding is calcium-dependent.
- cDNA cloning revealed p79 shares similarity with gastrin-binding protein and mitochondrial enzymes, while p46 is similar to mitochondrial ketoacyl-CoA thiolase.
- p79 localizes to the secretory pathway (ER, Golgi) and associates with calnexin; it forms a complex with FGF-2 and an MG-160-like protein in vivo.
Conclusions:
- A novel FGF-2 binding complex involving p150, p79, and p46 has been identified.
- Protein p79 plays a significant role in FGF-2 binding and cellular localization within the secretory pathway.
- The findings suggest a new mechanism for FGF-2 signaling potentially involving interactions within the endoplasmic reticulum and Golgi apparatus.