MOLECULAR ANALYSIS OF ETHYLENE OXIDE-INDUCED MUTATIONS AT THE HPRT LOCUS IN HUMAN-DIPLOID FIBROBLASTS

Ethylene oxide (EtO)-induced mutations in the hypoxanthine-guanine pho sphoribosyltransferase (HPRT) gene were characterized in 28 independen tly derived 6-thioguanine-resistant human diploid fibroblast clones us ing polymerase chain reaction-based techniques and Southern blot analy sis. Sequence analysis revealed one single base pair deletion and 13 b ase substitutions, nine of which were transversions: five AT --> TA, t hree GC --> TA and one GC --> CG. Four mutants were found to have GC - -> AT transitions. Seven of the point mutations caused splicing errors . Six occurred in splice site sequences and one created a new splice a cceptor site 16 bp upstream of exon 9. Three splice mutations were loc alized at the same site in the splice donor sequence of intron 8. Four teen mutants had large HPRT gene deletions. In seven mutants the entir e HPRT gene was deleted. The remaining deletion mutants had a truncate d HPRT gene, where one or several exons were lost. These results show that EtO induces many different kinds of HPRT mutations, among which a s many as 50% are large deletions.