Activation of N-ras in a human melanoma cell line.

Abstract

DNA isolated from cell line Mel Swift, a human melanoma cell line, transforms NIH3T3 cells. Southern blot analysis of DNA from secondary foci revealed conserved 8.8- and 7.8-kilobase EcoRI fragments which hybridized with a human repetitive sequence clone, blur 8. The activated transforming gene was identified as N-ras, and the 8.8-kilobase EcoRI fragment from a secondary transformant was cloned. Synthetic 17-mer oligonucleotides which spanned either the normal codon 61 (CAA) or a mutant codon 61 (AAA) were used for hybridization. Cloned N-ras from melanoma cell line Mel Swift hybridized to the mutant (AAA) oligonucleotide. From this we predicted a glutamine-to-lysine substitution in amino acid 61, a change confirmed by conventional sequencing of the first and second exons of N-ras from cell line Mel Swift. Transfection experiments showed that only those recombinant clones with the mutation in position 61 were biologically active.

Structural alteration in the MYB protooncogene and deletion within the gene encoding alpha-type protein kinase C in human melanoma cell lines.

“…Genetic studies of melanoma-prone kindreds indicate a highly penetrant, autosomal, dominant mode of inheritance for a dysplastic nevus syndrome/CMM-susceptibility gene with provisional linkage to the Rh locus on the short arm of chromosome 1 (7,8). Experimental transformation of mouse NIH 3T3 cells with human melanoma DNA has detected the sporadic activation of the oncogenes HRAS (9, 10), NRAS (10,11), and the RAS-related gene MEL (12,13). Cytogenetic studies of melanoma cell cultures have revealed nonrandom chromosomal alterations most frequently involving chromosomes 1, 6, and 7 (14-20).…”

N-ras mutations in human cutaneous melanoma from sun-exposed body sites.

“…These data imply that mutational activation of the N-rcas gene must have occurred during development of the primary tumor. Mutated ras genes, usually N-,-as, have been detected previously in melanoma metastases and cell lines but not in primary tumors (1,11,12,15). In one study, an N-ra-.s mutation was found to be present in one of five cell lines established from different metastases of one patient (1).…”

Understanding Melanoma Signaling Networks as the Basis for Molecular Targeted Therapy

“…Although MAPK activity in melanoma cells can arise through autocrine growth factor stimulation (Nesbit et al, 1999), N-cadherinbased homotypic cell-cell adhesion (Li et al, 2001), and melanoma cell--matrix adhesion, it is more commonly activated after the acquisition of an activating oncogenic mutation. The first such MAPK-activating mutation to be reported in melanoma was in NRAS (Padua et al, 1984(Padua et al, , 1985. Mutations in NRAS have since been identified in 15-20% of all melanomas, and are most commonly the result of the substitution from leucine to glutamine at position 61 (Brose et al, 2002;Davies et al, 2002).…”