Restoration of UV sensitivity in UV-resistant HeLa cells by antisense-mediated depletion of damaged DNA-binding protein 2 (DDB2).
Damaged DNA-binding activity comprises two major protein components, DDB1 and DDB2, which are implicated in the repair of ultraviolet (UV) radiation-induced DNA damage. The possible role of DDB2 as a determinant of cellular sensitivity to UV was investigated. The abundance of DDB2 in UV-resistant HeLa cell lines was increased compared with that in the parental UV-sensitive cells. Stable transfection of the resistant cells with DDB2 antisense cDNA resulted in marked depletion of DDB2 protein and restored cellular sensitivity to UV-induced apoptosis. Whereas the extent of UV-induced activation of apoptosis executioners, including DNA fragmentation factor, and caspase-3 were reduced in the UV-resistant cells compared with those apparent in the sensitive cells, depletion of DDB2 from the resistant cells restored the normal activation patterns for these proteins. In contrast, overexpressing DDB2 in DDB2-depleted cells with recombinant adenovirus, which carries ddb2 cDNA, markedly inhibited the extent of UV-induced activation of DNA fragmentation factor, and caspase-3. Interestingly, a mutated form of DDB2, which is defective in interacting with DDB1 and binding to UV-damaged DNA, also markedly inhibited the activation of apoptosis executioners. These results indicate that DDB2 is a modulator of UV-induced apoptosis, and that UV resistance can be overcome by inhibition of DDB2. The findings also suggest that modulation of UV-induced apoptosis by DDB2 may be independent of DNA repair.
Sun
Nian Kang
NK
Tumor Biology Laboratory, Department of Biochemistry, Chang Gung University, 333, ROC, Taoyuan, Taiwan.
Kamarajan
Pachiyappan
P
Huang
Haimei
H
Chao
Chuck C-K
CC
D017209:Apoptosis; D016373:DNA, Antisense / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000172:deficiency / Q000378:metabolism*; D004307:Dose-Response Relationship, Radiation; D006367:HeLa Cells; D006801:Humans; D014466:Ultraviolet Rays / Q000009:adverse effects*
C117669:DDB2 protein, human; D016373:DNA, Antisense; D004268:DNA-Binding Proteins