Involvement of nucleophosmin/B23 in the response of HeLa cells to UV irradiation.
The steady-state mRNA level of nucleophosmin/B23 in HeLa cells increased after UV irradiation. Nucleophosmin/B23 antisense transfection potentiated ultraviolet (UV)-induced cell killing. A block in G(2)/M phase, larger peak of apoptotic cells and higher caspase-3 in vitro activity were noted in nucleophosmin/B23 antisense-transfected cells compared with vector-transfected cells after UV treatment. Irradiated cells that received vector plasmid exhibited increased levels of [(3)H]thymidine incorporation due to DNA repair synthesis. In contrast, irradiated cells that received nucleophosmin/B23 antisense plasmid exhibited no such increase of [(3)H]thymidine incorporation, indicating inhibition of DNA repair. Cotransfection of cells with vector allowed repair of the damaged chloramphenicol acetyl transferase (CAT) reporter and rescue of CAT activity by host repair machinery. CAT activity in cells cotransfected with nucleophosmin/B23 antisense was less (<50%) than that of vector-transfected cells, indicating reduction of host nucleotide excision repair activity. Lower protein expressions of nucleophosmin/B23 and proliferating cell nuclear antigen (PCNA) were observed in nucleophosmin/B23 antisense-transfected cells compared with vector-transfected cells with or without UV treatment. Cotransfection of nucleophosmin/B23 antisense-transfected HeLa cells with PCNA construct made the cells less susceptible to UV-induced cell killing. Our results indicate that nucleophosmin/B23 correlates with PCNA and DNA repair capacity in cellular sensitivity to UV.
Wu
Ming H
MH
Cancer Biochemistry Laboratory, Department of Pharmacology, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-San, Tao-Yuan 333, Taiwan, Republic of China.
Chang
Jei H
JH
Chou
Chih C
CC
Yung
Benjamin Y M
BY
D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D004247:DNA / Q000528:radiation effects; D004260:DNA Repair; D004307:Dose-Response Relationship, Radiation; D005434:Flow Cytometry; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D008156:Luciferases / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D000090243:Nucleophosmin; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D010957:Plasmids / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D014466:Ultraviolet Rays
C000716967:NPM1 protein, human; D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense; D018809:Proliferating Cell Nuclear Antigen; D012333:RNA, Messenger; D000090243:Nucleophosmin; D012313:RNA; D004247:DNA; D008156:Luciferases; D015500:Chloramphenicol O-Acetyltransferase; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases