IDENTIFICATION OF AN AMPLIFIED GENE-CLUSTER IN GLIOMA INCLUDING 2 NOVEL AMPLIFIED GENES ISOLATED BY EXON TRAPPING

Gene amplification, which occurs in more than 50% of malignant gliomas , is considered to play a pivotal role in tumorigenesis. There are, ho wever, few studies aimed toward the isolation of novel genes from ampl ified sequences. Previously, we reported amplification of the protoonc ogene MET (hepatocyte growth factor receptor; 7q13) in more than 20% o f glioblastomas. For an approximate size estimation of the amplificati on unit we analyzed three glioblastomas all of which carried an amplif ied MET gene, by Southern blot analysis and/or competitive polymerase chain reaction using eight DNA markers. Although the extent of the amp lified domain varied, the close vicinity of the MET gene was the only region consistently amplified in these glioblastomas. A yeast artifici al chromosome (YAC) contig of 900 kb was refined spanning the amplifie d region flanking the MET gene, The YAC inserts were subcloned Into 59 cosmids, which were used for exon trapping, Eight sequences were iden tical to parts of the genes MET and CAPZA2 (human actin capping protei n alpha-subunit). Two newly identified exons and the CAPZA2 exons were amplified in tumor TX3095, which retains an amplified MET gene. The n ew exons were localized close to MET and CAPZA2, Characterization of t he clones, which were termed glioma-amplified sequence (GAS)7-1 and GA S7-2, showed an open reading frame and a different expression pattern in multiple human tissues. This study reports the identification of a cluster of amplified genes including two novel genes in a region ampli fied in more than 20% of glioblastomas.