[Pulmonary carcinogenesis susceptibility-associated single-nucleotide polymorphisms in K-ras intron 2 affect the...

The sequence of K-ras intron 2, which has been associated with lung tumor susceptibility in inbred mouse strains, was analyzed in susceptible strain GR and in resistant strains PT and UT. In the latter case, the intron had a tandem repeat of a 37-bp sequence with variant GC of its two single-nucleotide polymorphisms (SNPs), as earlier reported for resistant strains AKR, C57BL/c, and C3H/A. Strain GR did not differ in intron structure from susceptible strains A/He and ICR, having one copy of the 37-bp sequence with SNP variant CA. By gel retardation assay, a DNA probe corresponding to "susceptible" allele CA of K-ras region 278-307 formed an additional complex with nuclear proteins extracted from the lungs, as compared with probes corresponding to the "resistant" GC and "intermediate" CC alleles. With specific antibodies, the protein binding to the susceptible allele was identified as transcription factor GATA-6. Reverse transcription with subsequent multiplex PCR did not reveal a significant difference in K-ras expression for susceptible and resistant strains. The results suggest that SNPs of K-ras intron 2 do not affect the level of K-ras expression but do control the binding of GATA-6, which plays an important role in lung differentiation.