Novel electrophoretic protocol for collection of mutations in the lambda light chain immunoglobulin gene in a human...

Spontaneous and chemically induced mutation was examined in the lambda light chain immunoglobulin gene in a human B-lymphoblastoid cell strain (T5-1). The hemizygous lambda gene is a unique mutational target gene which codes for a protein that is both expressed on the cell membrane and secreted. Mutations in the lambda gene were detected by analysis of western blots of isoelectric focusing gel electrophoresis of T5-1 cell conditioned culture medium. None of 5,841 individual clones established from vehicle-exposed populations had detectable variations in the isoelectric banding pattern of the constitutively secreted lambda immunoglobulin protein. In contrast, 113 of 6,128 clonal populations established from N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-exposed populations exhibited stable variations in expression of the lambda immunoglobulin: isoelectric variants (n = 3) and non-secretors (n = 110). MNNG-induced mutations in the lambda gene, which resulted in lambda immunoglobulin proteins with altered isoelectric points (pIs), occurred at a frequency of no less than 4.9 x 10(-4) mutations/cell, indicating the mature rearranged lambda immunoglobulin gene is comparably sensitive to carcinogen induced mutation as other human autosomal target genes. Approximately one-half of the MNNG-induced non-secretor mutant clones lacked lambda mRNA while one-half maintained constitutive transcription and expression of the lambda immunoglobulin on the cell surface, demonstrating that carcinogen damage interdicted gene function at multiple points.