ABSTRACT
PURPOSE: To gain insight on transcriptional repression of Topo II a in HL-60 cells arrested to G2/M and M phase, the levels of Topo IIa mRNA and the binding activity of ATF have been investigated with Northern blot hybridization and DNA mobility shift assay, respectively. MATERIALS AND METHODS: HL-60 cells were grown in RPMI 1640 medium supplemented with 10% heat-mactivated fetal bovine serum and antibiotics in a humidified 5% CO2 at 37C degree. Total RNA was prepared by a modification of the method of Karlinsey et al. Northern blot hybridization was performed by the method of Virca et al. A Xho I-Mlu I fragment of phTOP2 was used as probe for Northern blot analysis of Topo II a mRNA. DNA mobility shift assay was performed by the method of Lim et al. End labeled DNA oligomer (upper strand, 5-TCTCCGCTATGACGCCGAGTGGTG-3) for ATF binding activity was mixed with nuclear extracts in a 20 pl reaction volume containing 60 mM KC1, 12 mM HEPES, pH 7.9, 5 mM MgCl2, 0.2 mM EDTA, 0.2 mM DTT, 12% glycerol, and 2 ug of poly [dI-dC]. RESULTS: HL-60 cells were arrested at G2/M phase and M phase after taxol or nocodazole treatment. The levels of Topo II a mRNA were reduced at 24 hours after exposure with nocodazole or taxol but the unknotting activities were not changed. DNA mobility shift assay using oligonucleotide containing the ATF binding site showed that ATF binding activity was reduced after pretreatment of nododazole or taxol. CONCLUSIONS: These results suggest that the reduction of ATF binding activity may be important to transcriptional repression of Topo II a gene by nocodazole and taxol in HL- 60 cells.
Subject(s)
Humans , Anti-Bacterial Agents , Binding Sites , Blotting, Northern , Cell Division , DNA Topoisomerases, Type I , DNA Topoisomerases, Type II , DNA , Edetic Acid , Electrophoretic Mobility Shift Assay , Genes, vif , Glycerol , HEPES , HL-60 Cells , Hydrogen-Ion Concentration , Magnesium Chloride , Nocodazole , Paclitaxel , Repression, Psychology , RNA , RNA, MessengerABSTRACT
Uterine didelphys is a congenital malformation characterized by the presence of two separated hemiuteri andhemivaginas, due to lack of midfusion of the Mullerian ducts. We report a case of UD-BHRA(uterine didelphys withblind hemivagina and ipsilateral renal agenesis), a rare type of uterine didelphys charaterized by symptomaticunilateral hematocolpos due to blind hemivagina after menarche and ipsilateral renal agenesis. The MRI findings in22-year-old woman with bilateral ovarian tumors demonstrated two separated uterine horns and cervical and vaginalcanals, with left hematocolpos and left renal agenesis.