ENHANCED REPAIR ENDONUCLEASE ACTIVITIES FROM RADIATION-ARRESTED G(2) PHASE MAMMALIAN-CELLS

HeLa cells arrested in Gz phase 22 h after receiving 11.5 Gy gamma-rad iation contained 3.6-fold more EDTA-resistant DNA repair endonuclease activity than unirradiated cells. Enzyme activity was determined by me asuring the release of fragments from an irradiated repetitive alpha D NA substrate or from synthetic substrates containing a single modified base, 8-oxoguanine (8-oxo-G), a major radiation product. It appeared that the radiation-induced enhanced repair activity in some cells migh t be a feature of radiation-induced G(2) arrest. Indeed, unirradiated G(2) HeLa cells that had been synchronized by double thymidine block c ontained 3-7-fold more endonuclease activity than G(1) or S-phase cell s. Similarly, two of four other cell lines tested exhibited elevated r epair endonuclease activity in G(2). However, all six cell lines teste d exhibited radiation-enhanced repair endonuclease activity. Therefore , the underlying mechanism for radiation enhancement of enzyme activit y remains to be clarified and does not seem to be completely accounted for as a consequence of G(2) arrest. The results showed different sub strate specificities among cell lines as well as differences during th e cell cycle of individual cell lines. Repair endonuclease activity fr om all cell lines which we have tested were associated with 60-70 kDa proteins from Superose 12 columns. Since reports from other laboratori es have described several different DNA repair activities in 50-70 kDa Superose 12 fractions, it seems possible that the DNA repair enzymes may be associated in a repairosome structure.