Caffeine enhancement of alkylating agent-induced injury in barley: Its connection to DNA single-strand breaks and their repair

Abstract

Non-germinating barley seeds were treated with N -methyl- N -nitrosourea (MNU), methyl methanesulphonate (MMS) or ethyl methanesulphonate (EMS) and then exposed either to non-toxic caffeine doses ( i.e. doses having no biological effect and inducing no DNA single-strand breaks when applied alone) or to toxic doses ( i.e. doses reducing seed germination and seedling height but inducing no DNA single-strand breaks). Non-toxic doses of caffeine, applied to MNU, MMS and EMS treated seeds enhanced the mutagen-induced reduction of germination and seedling height but did not disturb the recovery from this damage, achieved by storage of the seeds at 30% water content. At the DNA level, these doses of caffeine did not increase the amount of mutagen-induced single-strand breaks in the treated (preformed) DNA and did not inhibit repair of mutagen-induced single-strand breaks during seed storage. However, non-toxic doses of caffeine affected the size of DNA strands, newly synthesized during 3 days of seed germination. Whereas the size of DNA strands after MMS and EMS treatment approached the size of the newly synthesized DNA in the controls, the DNA strands from treatment with MMS or EMS plus non-toxic caffeine doses were considerably smaller. Thus, it seems likely that caffeine, although applied before the DNA-synthesis, enhanced the biological damage by acting in the S-phase and inhibiting the repair of DNA damage in the newly synthesized DNA strands. Interaction of toxic caffeine doses with MNU led both to the potentiation of the mutagen-induced reduction of germination and seedling height, and to the inhibition of recovery from this damage during seed storage. On the DNA level. even toxic doses of caffeine did not increase the amount of MNU-induced single-strand breaks in the preformed DNA but inhibited completely the repair of these breaks during the seed storage.