INTERACTIONS BETWEEN THE FLK-1 RECEPTOR, VASCULAR ENDOTHELIAL GROWTH-FACTOR, AND CELL-SURFACE PROTEOGLYCAN IDENTIFIED WITH A SOLUBLE RECEPTOR REAGENT
INTERACTIONS BETWEEN THE FLK-1 RECEPTOR, VASCULAR ENDOTHELIAL GROWTH-FACTOR, AND CELL-SURFACE PROTEOGLYCAN IDENTIFIED WITH A SOLUBLE RECEPTOR REAGENT
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DOI：
10.3109/08977199509003208
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发表时间：
1995-01-01
期刊：
影响因子：
1.8
通讯作者：
FLANAGAN, JG
中科院分区：
文献类型：
--
作者：
CHIANG, MK;FLANAGAN, JG
胎儿肝激酶-L(Flk-1)是一种跨膜酪氨酸激酶，存在于血管内皮细胞和大量富含造血祖细胞的细胞中。为了研究Flk-1与潜在配体的相互作用，我们将受体胞外区与碱性磷酸酶(AP)标签进行了基因融合。通过与Flk1-AP融合蛋白的免疫共沉淀，在许多间充质细胞系的培养上清液中鉴定出Flk-1的可溶性配体。N-末端测序结果表明该多肽为血管内皮生长因子(VEGF)。因此，受体-AP融合蛋白可以用于识别可溶性配体和跨膜配体，因此这种方法很可能广泛适用于许多类型的孤儿受体。Flk1-AP可溶性受体也能与细胞表面结合，表现出两种不同亲和力的明显结合部位。通过将血管内皮生长因子表达载体导入细胞，可以重建这种相互作用。这些结果表明，出现在细胞表面的血管内皮生长因子可以与Flk-1受体结合，并可能介导细胞与细胞的直接相互作用。Flk1-AP融合蛋白还能与肝素结合，这意味着Flk-1受体与配体的结合可能涉及Flk-1受体、血管内皮生长因子和肝素样细胞表面蛋白多糖之间的三向相互作用。
Fetal Liver kinase-l (Flk-1) is a transmembrane tyrosine kinase that was identified in endothelial cells and populations of cells enriched in hematopoietic progenitors. To characterize the interaction of Flk-1 with potential ligands the receptor extracellular domain was genetically fused to an alkaline phosphatase (AP) tag. A soluble ligand for Flk-1 was identified in the supernatants of numerous mesenchymal cell lines by co-immunoprecipitation with the Flk1-AP fusion protein. This polypeptide was shown by N-terminal sequencing to be vascular endothelial growth factor (VEGF). Receptor-AP fusion proteins can thus be used to identify soluble ligands as well as transmembrane ligands, and this approach is therefore likely to be widely applicable to many types of orphan receptor. The Flk1-AP soluble receptor was also found to bind to cell surfaces, showing two apparent classes of binding site with different affinities. This interaction could be reconstructed by introducing a VEGF expression plasmid into cells. These results indicate that VEGF presented at the cell surface can bind to the Flk-1 receptor, and could mediate a direct cell-cell interaction. The Flk1-AP fusion protein was also found to bind heparin, implying that ligand binding by the Flk-1 receptor may involve a three way interaction between the Flk-1 receptor, VEGF, and heparin-like cell surface proteoglycans.