BIOCHEMICAL-CHARACTERIZATION OF 2 ISOFORMS OF FLT4, A VEGF RECEPTOR-RELATED TYROSINE KINASE

The FLT4 gene encodes a tyrosine kinase receptor related to the two id entified receptors for vascular endothelial growth factor (VEGF), FLT1 and FLK1/KDR. Two isoforms of FLT4, differing by their C-terminal end s, have been identified. The long form has 65 additional amino acid re sidues. We have shown that FLT4 is a highly glycosylated, relatively s table, cell surface associated kinase of approximately 180 kDa. In ord er to study the signal transduction molecules associated with the FLT4 pathway, and in the absence of a known ligand, we constructed two chi meric molecules (FF4S and FF4L) made of the extracellular region of th e CSF1 receptor (Fms gene product) and of the transmembrane and intrac ellular regions of either form of FLT4. These two chimeric forms were expressed in Rat 2 transfectants. We assayed the ligand-induced capaci ty of the FF4 short and long forms to sustain growth of Rat 2 cells in semisolid medium. In a soft agar assay, only the long form was able t o induce the growth of Rat 2 cells upon ligand treatment. The two form s of FLT4 therefore have different functional capacities. We looked fo r association and/or phosphorylation of phospholipase C gamma (PLC gam ma) and phosphatidylinositol-3'-phosphate (PI3K), after stimulation of the FF4 molecules by CSF1. Finally, we have studied the expression of the Flt4 gene in mouse embryos and in the adult by in situ hybridizat ion. Flt4 transcripts were found at day 12.5 post-coitum and thereafte r, including the adult mouse, predominantly in the pericardium, pleura l membranes and in the lung.