Development of Zidovudine (AZT) Resistance in Jurkat T Cells Is Associated with Decreased Expression of the Thymidine Kinase (TK) Gene and Hypermethylation of the 5' End of Human TK Gene
作者:
Shijuan Wu,
Xiping Liu,
Michelle Solorzano,
Richard Kwock,
Vassilios Avramis,
期刊:
Journal of Acquired Immune Deficiency Syndromes and Human Retrovirology
(OVID Available online 1995)
卷期:
Volume 8,
issue 1
页码: 1-9
ISSN:1077-9450
年代: 1995
出版商: OVID
关键词: AZT;Thymidine kinase;Reverse transcriptase.
数据来源: OVID
摘要:
The T-cell line Jurkat E6–1 was rendered resistant to zidovudine (AZT) in vitro by exposure to low but gradually increased concentrations of the drug. Biochemical pharmacology studies of [3H]AZT in the AZT-resistant T-cell lines showed a significant reduction of AZT phosphorylation to the mono-, di-, and triphosphate anabolites. Peripheral blood mononuclear cells (PBMCs) from pediatric patients with human immunodeficiency virus type 1 (HIV-1) infection showed a similar pattern of decreased AZT anabolism. Enzymatic studies with purified thymidine kinase (TK) preparations from these cell lines showed a gradual decline inVmaxrelated to their level of resistance to AZT. The Jurkat/AZT-20 and Jurkat/AZT-100 cells were studied in greater detail with reverse transcriptase/polymerase chain reaction (RT/PCR) cloned probes to determine possible molecular mechanisms of resistance to AZT. TK mRNA was significantly decreased (∼5− to 10-fold) in the AZT-resistant T-cell lines. Southern blot analyses indicated that there were no major rearrangements or deletions of theTKgene, but the 5‘ end of the gene in the AZT-resistant cells is highly methylated when compared to wild-type cells. No apparent differences were seen in thymidylate kinase (dTMPk) mRNA levels in the same T-cell lines. Thus the decreased expression of TK mRNA and resultant TK enzymatic activity is responsible for the observed reduction in the AZT anabolism in the resistant T-cell lines. Decreased T-cell TK activity could allow wild-type, AZT-sensitive HIV-1 to replicate in the presence of subinhibitory AZT triphosphate (AZT-TP) cellular concentrations enabling a genetic variant with drug resistance to emerge and outgrow the AZT-sensitive, wild-type virus.
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