利用小干扰RNA对人脱氧尿苷三磷酸酶进行调控

Modulation of human dUTPase using small interfering RNA.

作者信息

Studebaker A W, Lafuse W P, Kloesel R, Williams M V

机构信息

Department of Molecular Virology, Immunology and Medical Genetics, The Ohio State University, Columbus, OH 43210, USA.

出版信息

Biochem Biophys Res Commun. 2005 Feb 4;327(1):306-10. doi: 10.1016/j.bbrc.2004.12.021.

DOI:10.1016/j.bbrc.2004.12.021

PMID:15629463

Abstract

Deoxyuridine triphosphate nucleotidohydrolase (dUTPase) is responsible for maintaining low intracellular levels of dUTP, thus preventing the incorporation of dUTP into DNA. A 21 bp double-stranded RNA molecule (siRNAdUT3) targeted against motif 3 of human dUTPase resulted in a time- and dose-dependent decrease in dUTPase activity in transfected cells. dUTPase activity was reduced approximately 95+/-5% in all cell lines tested 48 h after transfection with 2 microg siRNAdUT3 and it was maintained at this decreased level for at least 72 h. Down-regulation of dUTPase resulted in a significant increase in intracellular dUTP and a decreased proliferation of the transfected cells. Therefore, we conclude that dUTPase activity/expression can be down-regulated using siRNA specifically targeted to dUTPase mRNA and that this approach can be used to elucidate the role of dUTPase in DNA metabolism, as well as, to determine whether dUTPase is a valid target for drug development.

摘要

脱氧尿苷三磷酸核苷酸水解酶（dUTPase）负责维持细胞内低水平的dUTP，从而防止dUTP掺入DNA。一个针对人类dUTPase基序3的21 bp双链RNA分子（siRNAdUT3）导致转染细胞中dUTPase活性呈时间和剂量依赖性降低。用2 μg siRNAdUT3转染后48小时，所有测试细胞系中的dUTPase活性均降低了约95±5%，并在该降低水平维持至少72小时。dUTPase的下调导致细胞内dUTP显著增加，转染细胞的增殖减少。因此，我们得出结论，使用特异性靶向dUTPase mRNA的siRNA可以下调dUTPase活性/表达，并且这种方法可用于阐明dUTPase在DNA代谢中的作用，以及确定dUTPase是否是药物开发的有效靶点。