Cell cycle progression in denV-transfected murine fibroblasts exposed to ultraviolet radiation.
Repair-proficient murine fibroblasts transfected with the denV gene of bacteriophage T4 repaired 70-80% of pyrimidine dimers within 24 h after exposure to 150 J/m2 ultraviolet radiation (UVR) from an FS-40 sunlamp. Under the same conditions, control cells repaired only about 20% of UVR-induced pyrimidine dimers. After UVR exposure, both control and denV-transfected cells exhibited some degree of DNA-synthesis inhibition, as determined by flow cytometric analysis of cell-cycle kinetics in propidium iodide-stained cells. DenV-transfected cells had a longer and more profound S phase arrest than control cells, but both control and denV-transfected cells had largely recovered from UVR effects on cell-cycle kinetics by 48 h after UVR exposure. Inhibition of DNA synthesis by UVR was also measured by determining post-UVR incorporation of bromodeoxyuridine (BrdU). The amount of BrdU incorporated was quantitated by determining with flow cytometry the quenching of Hoechst dye 33342 by BrdU incorporated in cellular DNA. DenV-transfected cells showed more marked inhibition of BrdU incorporation after low fluences of UVR than control cells. Differences between denV-transfected and control cells in cell-cycle kinetics following UVR exposure may be related to differences in mechanisms of repair when excision repair of pyrimidine dimers is initiated by endonuclease V instead of cellular repair enzymes.
Kusewitt
D F
DF
Center for Photomedicine, Lovelace Medical Foundation, Albuquerque, NM 87108.
Budge
C L
CL
Nolla
H A
HA
Edwards
B S
BS
Ley
R D
RD
D000818:Animals; D001973:Bromodeoxyuridine / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects*; D004260:DNA Repair; D043224:Deoxyribonuclease (Pyrimidine Dimer); D004706:Endodeoxyribonucleases / Q000235:genetics*; D005434:Flow Cytometry; D005814:Genes, Viral; D066298:In Vitro Techniques; D051379:Mice; D011740:Pyrimidine Dimers / Q000378:metabolism; D014162:Transfection; D014466:Ultraviolet Rays; D015678:Viral Structural Proteins / Q000235:genetics
D011740:Pyrimidine Dimers; D015678:Viral Structural Proteins; D004706:Endodeoxyribonucleases; D043224:Deoxyribonuclease (Pyrimidine Dimer); D001973:Bromodeoxyuridine