c-met is amplified but not mutated in a cell line with an activated met tyrosine kinase

The putative tyrosi ne kinase receptor encoded by the oncogene c-met is activated (tyrosine-phosphorylated in vivo) in the human gastric carcinoma cell line GTL-16. The corresponding gene is amplified and over-expressed. In this study we show that c-met is part of an amplification unit measuring more than 3000 kb. The multiple copies of the amplicon are located on a novel chromosome different from chromosome 7. We have previously shown that the c-met protein present in GTL-16 cells is indistinguishable from that found in other cells. Kinase activation could be due to over-expression of the normal c-met protein or to the presence of activating mutation(s). To verify the primary structure of the c-met protein in GTL-16 cells we sequenced a series of overlapping cDNAs obtained from GTL-16 cell RNA by reverse transcription and polymerase chain reaction. Two differences were found in the c-met coding region with respect to the published human c-met cDNA: (1) the lack of 54 nucleotides corresponding to a stretch of 18 amino acids located in the extracellular domain of the receptor, and (2) the substitution of the codon specifying alanine 1209 (located in the tyrosine kinase domain) with one coding for glycine. However, we also obtained cDNAs identical to that just described from a number of control cell lines. These results suggest: (1) that the present c-met cDNA presumably reflects the sequence of the most abundant transcript in several cell types, and (2) that overexpression of the normal c-met protein, alone or in combination with an autocrine loop, is most probably responsible for the activation of the c-met kinase in GTL-16 cells.