Coamplification in tumors of KRAS2, type 2 inositol 1,4,5 triphosphate receptor gene, and a novel human gene, KRAG.
Analysis of a region of DNA, coamplified in tumors with KRAS2, resulted in the identification of the human homologue of the mouse KRAG gene. The gene was widely expressed in a range of cell lines, tumors, and normal tissue and demonstrated a high degree of alternate splicing. A human KRAG cDNA sequence, with a structure similar to that encoded by the amplified gene in mouse Y1 adrenal carcinoma cells, was isolated by RT-PCR. The predicted amino acid similarity between the two sequences was 91%, and hydrophobicity plots suggested a structure closely resembling that of transmembrane 4 superfamily members. Identification of a PCR-based restriction fragment length polymorphism confirmed biallelic expression of KRAG but suggested allele-specific splicing differences in tumors. Northern analysis of mRNA derived from a range of tissues suggested high level expression in muscle and confirmed alternate splicing. To facilitate the analysis of exon junctions, a YAC clone encoding the genomic sequence was identified. This allowed the localization of KRAG to human chromosome 12p11.2. Isolation of one end of this nonchimeric clone demonstrated a perfect match with a 247-bp sequence within the 3' untranslated region of the type 2 1,4, 5-inositol triphosphate receptor gene. Multiplex PCR confirmed the inclusion of both genes in the KRAS2 amplicon in human malignancy, suggesting that either may contribute to the malignant phenotype.
Heighway
J
J
CRC Department of Cancer Genetics, Paterson Institute for Cancer Research, Christie Hospital (NHS) Trust, Wilmslow Road, Manchester, M20 9BX, United Kingdom.
Betticher
D C
DC
Hoban
P R
PR
Altermatt
H J
HJ
Cowen
R
R
D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015220:Calcium Channels / Q000235:genetics*; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000473:pathology; D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000473:pathology; D002352:Carrier Proteins; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D002881:Chromosomes, Human, Pair 12 / Q000235:genetics* / Q000648:ultrastructure; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics*; D005784:Gene Amplification; D015972:Gene Expression Regulation, Neoplastic; D011905:Genes, ras; D006801:Humans; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008175:Lung Neoplasms / Q000235:genetics / Q000473:pathology; D008565:Membrane Proteins / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D009369:Neoplasms / Q000235:genetics*; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D014407:Tumor Cells, Cultured
D015220:Calcium Channels; D002352:Carrier Proteins; D018076:DNA, Complementary; D004273:DNA, Neoplasm; C506926:ITPR1 protein, human; C506927:ITPR2 protein, human; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D018160:Receptors, Cytoplasmic and Nuclear; C109646:SSPN protein, human; C491632:Sspn protein, mouse