Chromosome-mediated transfer of murine alleles for hypoxanthine-guanine phosphoribosyl transferase (HGPRT) and ouabain resistance into human cell lines

Abstract

Genetic drug-resistance markers were transferred via purified metaphase chromosomes from mouse L cells into the human fibrosarcoma line HT1080 and HeLa S3 cells. Interspecific chromosome-mediated transfer of hypoxanthine-guanine phosphoribosyl transferase (HGPRT; EC 2.4.2.8) from mouse L cells into HGPRT- HT1080 cells occurred at a frequency of approximately 1 x 10(-7). The presence of the mouse allele for HGPRT in transferent isolates was confirmed by isoelectric focusing. Transfer of ouabain resistance from mouse L cells to HT1080 and HeLa S3 cells occurred at an average frequency of approximately 4 x 10(-7). Expression of the mouse trait in transferent isolates was confirmed by their ability to withstand doses of ouabain which would be lethal to spontaneous ouabain-resistant mutants of the human cells but not to mouse L cells, ouabain-resistant transferents of human cells showed 10(4)-to greater than 10(5)-fold enhanced drug resistance, characteristic of either wild-type or mutant alleles, respectively, from ouabain-resistant donor L cells. Unstable expression of the transferred phenotypes in the absence of selection was seen in some isolates, but expression was lost at slow rates.

Functional expression of the genomic DNA sequences encoding mouse Na,K-ATPase alpha 1 gene by cotransfection of overlapping genomic DNA segments.

“…Rodent and primate cell lines show extreme differences in ouabain sensitivity, a feature which has been exploited extensively in somatic cell genetics (1,16). The transfer of ouabain resistance to ouabain-sensitive cells has been correlated with the presence of the rodent Na,KATPase al subunit gene (6,14). Gene transfer experiments using mouse and rat al subunit cDNAs have also provided evidence that it is the expression of the rodent al subunit that mediates ouabain resistance (5,11,18).…”

Chromosome-mediated transfer of the murine Na,K-ATPase alpha subunit confers ouabain resistance.

“…By using somatic cell hybrids, ouabain resistance has been assigned to mouse chromosome 3 (5). Chromosomemediated transfer of the murine ouabain resistance phenotype to human cell lines has also been achieved (8). However, the molecular basis for the species-specific difference in ouabain sensitivity has not been clearly explained.…”