REPAIR OF RIBOSOMAL-RNA GENES IN HAMSTER-CELLS AFTER UV IRRADIATION, OR TREATMENT WITH CISPATIN OR ALKYLATING-AGENTS

We have measured the DNA damage formation and repair in the ribosomal and the dihydrofolate reductase (DHFR) genes after treatment of hamste r cells with different types of DNA damaging agents. In mammalian cell s, the ribosomal DNA (rDNA) is transcribed by RNA polymerase I, wherea s the DHFR is transcribed by RNA polymerase II. Cells were treated wit h agents that induce different types of lesions, and that are known to be repaired via different pathways. We used UV (254 nm) irradiation, treatment with cisplatin and treatment with the alkylating agents nitr ogen mustard (HN2) and methyl methanesulphonate (MMS). UV induced pyri midine dimers were detected with the enzyme T4 endonuclease V, which c reates nicks at the dimer sites; the breaks are then resolved and iden tified by denaturing electrophoresis and Southern blot. Intrastrand ad ducts formed by the alkylating agents HN2 and MMS were quantitated by generating strand breaks at abasic sites after neutral depurination. I nterstrand crosslinks (ICL) formed by HN2 and cisplatin were detected by a denaturation-reannealing reaction before neutral agarose gel-elec trophoresis. We find that the repair of the pyrimidine dimers is signi ficantly less efficient in the RNA polymerase I transcribed rDNA genes than in RNA polymerase II transcribed DHFR gene at 8 and 24 h after i rradiation. ICL and intrastrand adducts induced by HN2 are also remove d more slowly from the rDNA than from the DHFR gene. In contrast, MMS induced intrastrand adducts and cisplatin induced ICL are repaired equ ally efficiently in the RNA polymerase I and RNA polymerase II transcr ibed genes. We conclude that for some types of DNA damage, there is le ss repair in the ribosomal genes than in the DHFR; but for other DNA l esions there is no difference. The difference in repair efficiency bet ween the rDNA and the DHFR genes may reflect the different RNA polymer ases involved in their transcription. It may, however, alternatively, reflect the different nuclear localization of these genes.