The second intron of the K-ras gene contains regulatory elements associated with mouse lung tumor susceptibility

We have previously demonstrated the preferential activation of the K-ras gene from the susceptible A/J parent in lung tumors from F1 mouse hybrids. In the present study, the mechanism of this observation is further investigated. Higher levels of expression of A/J K-ras allele were detected in lung adenomas (30 of 30) from the C3A mouse. In addition, three K-ras alleles, designated as susceptible (Ks), intermediate (Ki), or resistant (Kr), were identified by sequence analysis of the second intron of the K-ras gene from 32 strains of mice. These K-ras alleles are associated with differences in mouse lung tumor susceptibility. All Kr alleles have a tandem 37-bp direct repeat (nt 282-355) in the second intron of the K-ras gene. Ks and Ki alleles have only one copy of the 37-bp sequence (nt 282-318). Ks strains have three base variations at nt 288, 296, and 494, and Ki strains have two base variations at nt 288 and 494 in the second intron of the K-ras gene. Differential protein-binding patterns were observed in gel-mobility-shift experiments between the duplicated 37-bp sequence of the Kr allele and the single 37-bp sequence of the Ks and Ki alleles. DNase I footprinting assay revealed protein binding sites in the second intron of the K-ras gene that correspond to the tandem repeat sequences. Our data suggest that higher expression of the A/J allele relative to C3H allele may be responsible for the allele-specific activation of the K-ras gene in lung tumors from F1 hybrid mice.