Natural antisense (rTSα) RNA induces site-specific cleavage of thymidylate synthase mRNA
Natural antisense (rTSα) RNA induces site-specific cleavage of thymidylate synthase mRNA
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DOI：
10.1016/s0925-4439(02)00081-9
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发表时间：
2002-07-18
影响因子：
6.2
通讯作者：
Dolnick, BJ
中科院分区：
文献类型：
--
作者：
Chu, JX;Dolnick, BJ
The 3' untranslated region (UTR) of rTSalpha RNA is complementary (i.e., antisense) to human thymidylate synthase (TS) RNA. When HEp2 cells (human epidermoid carcinoma) progressed from late-log to plateau phase growth, ribonuclease protection assay (RPA) revealed an inverse correlation between the levels of rTSalpha RNA and TS mRNA, suggesting a possible effect of rTSalpha RNA on TS mRNA levels. HEp2 cells expressing a Tet-On transactivator were transiently co-transfected with pHook-1 and a construct containing rTSalpha (protein and antisense RNA), rTSalpha-3' (rTSalpha protein only), rTSalpha-3' (antisense RNA-luciferase) or luciferase. Transfected cells were selected and evaluated for the effects of induced transgene expression on TS mRNA. Induced expression of transfected rTSalpha or rTSalpha-3 but not rTSalpha-3' or luciferase, resulted in decreased TS mRNA levels as measured by RPA. These results demonstrated that the antisense region of rTSalpha RNA is necessary and sufficient for this down-regulation of TS mRNA. RPA for TS mRNA also showed the enhanced appearance of two partial-length protected fragments in rTSalpha or rTSalpha-3' transfected cells. RPA stringency evaluations and primer extension assays indicated that TS mRNA is cleaved in vivo in a site-specific manner. These data demonstrate that rTS gene expression likely plays a role in down-regulating TS through a natural RNA-based antisense mechanism. (C) 2002 Elsevier Science B.V All rights reserved.
