Double-strand break repair machinery is sensitive to UV radiation.
The precision of the repair of linearized plasmid DNA was analyzed using a nonsense mutation inactivated beta-glucuronidase (uidA) marker gene delivered to Nicotiana plumbaginifolia protoplasts and Nicotiana tabacum leaves. The reversions at the stop-codon allowed the reactivation of the marker gene. Here we report that irradiation of plant protoplasts or plant tissue prior to the delivery of the DNA repair substrate significantly potentiated the reversion frequency leading to a two to fourfold increase over the non-irradiated samples. The increase in reversion frequency was highest upon the delivery of the linear substrates, suggesting increased sensitivity of the double-strand break (DSB) repair apparatus to UV-C. Moreover, the most significant UV irradiation effect was observed in plasmids linearized in close proximity to the stop codon. The higher reversion frequency in UV-treated samples was apparently due to the involvement of free radicals as pretreatment of irradiated tissue with radical scavenging enzyme N-acetyl-l-cysteine abolished the effect of UV-C. We discuss the UV-sensitivity of various repair enzymes as well as possible mechanisms of involvement of error-prone polymerases in processing of DSBs.
Ilnytskyy
Yaroslav
Y
Department of Biological Sciences, University of Lethbridge, Lethbridge, AB, Canada T1K 3M4.
Yao
Youli
Y
Kovalchuk
Igor
I
D001483:Base Sequence; D018389:Codon, Nonsense / Q000235:genetics; D004249:DNA Damage; D004260:DNA Repair / Q000528:radiation effects*; D005609:Free Radicals / Q000378:metabolism; D005786:Gene Expression Regulation; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010957:Plasmids / Q000737:chemistry / Q000235:genetics / Q000528:radiation effects; D011523:Protoplasts / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D014026:Nicotiana / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism / Q000528:radiation effects*; D014466:Ultraviolet Rays
D018389:Codon, Nonsense; D005609:Free Radicals; D005966:Glucuronidase