Characterization of a UV-damage recognition factor in vitro that is associated with UV resistance in HeLa cells.
We have previously reported a cisplatin-selected HeLa cell line showing cross-resistance to ultraviolet (UV) radiation and overexpression of UV-damage recognition factors (Chao et al., Mol. Cell. Biol., 11, 2075-2080, 1991). Here, we further characterize a UV-damage recognition factor in vitro using a gel mobility shift assay. The results indicate that the damage-recognition factor is (i) localized mostly in the nucleus, (ii) protease-sensitive, (iii) RNA-independent, (iv) active in a wide range of ionic strengths (50-400 mM NaCl), (v) with a high affinity for UV-damaged DNA (50-fold molar excess competitor causes 50% recognition loss), and (vi) resistant to agents and that modify protein conformation (urea and NP-40), but slightly sensitive to CaCl2. The significance of the identified UV-damage recognition factor in the sensitivity or resistance of cells to UV is also discussed.
Chao
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Department of Biochemistry, Chang Gung Medical College, Taoyuan, Taiwan.
D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D002945:Cisplatin / Q000494:pharmacology; D004247:DNA / Q000528:radiation effects; D004249:DNA Damage / Q000235:genetics*; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000187:drug effects / Q000378:metabolism*; D004558:Electric Stimulation; D006367:HeLa Cells / Q000528:radiation effects*; D006801:Humans; D009687:Nuclear Proteins / Q000187:drug effects / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011487:Protein Conformation / Q000187:drug effects; D014466:Ultraviolet Rays
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D002945:Cisplatin