Endogenous Retroviral env Genes after N-Methyl-N′-nitro-N-nitrosoguanidine Treatment of Mouse Tumor Cells: Stable DNA Amplification and Rearrangement

Abstract

Immunogenic tumor variant clones derived by N-methyl-N′-nitro-N-nitrosoguanidine treatment of Eb lymphoma cells showed structurally altered gp70 env proteins at the cell surface. To further investigate this observation we screened for complementary DNA clones encoding gp70 antigens from a λ gt11 expression library constructed from mRNA of a mutant cell clone. Using gp70-specific antibodies, a total of 10 complementary DNA clones were identified and analyzed. DNA-sequence analysis revealed the presence of both xeno/or mink cell focus-forming-type (clones 1, 10) and endogenous ecotropic gp70 (clones 2, 3, 4). Southern blot experiments using clone 2 as a probe detected new restriction fragments and DNA amplification with high copy number in the DNAs of N-methyl-N′-nitro-N-nitrosoguanidine-treated but not in parental or 5′-azacytidine-treated control cells even after > 120 days in tissue culture. No alterations in the restriction fragment pattern of the genomic DNAs could be detected using H-2K-, β-actin-, or dehydrofolate-reductasespecific gene probes. The DNA amplification of specific gp70 sequences may be related to the enhanced expression of a subset of gp70 in mutant cells that was previously found to induce syngeneic antibody production.