Analysis of the distribution of DNA repair patches in the DNA-nuclear matrix complex from human cells.
The distribution of ultraviolet-induced repair patches along DNA loops attached to the nuclear matrix, was investigated by digestion with DNA-degrading enzymes and neutral sucrose gradient centrifugation. When DNA was gradually removed by DNAase 1, pulse label incorporated by ultraviolet-irradiated cells during 10 min in the presence of hydroxyurea or hydroxyurea/arabinosylcytosine showed similar degradation kinetics as prelabelled DNA. No preferential association of pulse label with the nuclear matrix was observed, neither within 30 min nor 13 h after irradiation. When the pulse label was incorporated by replicative synthesis under the same conditions, a preferential association of newly-synthesized DNA with the nuclear matrix was observed. Single-strand specific digestion with nuclease S1 of nuclear lysates from ultraviolet-irradiated cells, pulse labelled in the presence of hydroxyurea/arabinosylcytosine, caused a release of about 70% of the prelabelled DNA and 90% of the pulse-labelled DNA from the rapidly sedimenting material in sucrose gradients. The results suggest no specific involvement of the nuclear matrix in repair synthesis, a random distribution of repair patches along the DNA loops, and simultaneously multiple incision events per DNA loop.
D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism / Q000528:radiation effects*; D004260:DNA Repair; D004261:DNA Replication / Q000528:radiation effects; D004720:Endonucleases; D006801:Humans; D007700:Kinetics; D015719:Single-Strand Specific DNA and RNA Endonucleases; D014466:Ultraviolet Rays
D004720:Endonucleases; D015719:Single-Strand Specific DNA and RNA Endonucleases