Targeting RNA for degradation with a (2'-5')oligoadenylate-antisense chimera.
Antisense oligonucleotides hold considerable promise both as research tools for inhibiting gene expression and as agents for the treatment of a myriad of human diseases. However, targeted destruction of RNA has been difficult to achieve in a versatile, efficient, and reliable manner. We have developed an effective strategy for cleaving unique RNA sequences with 2-5A-dependent RNase, an endoribonuclease that mediates inhibitory effects of interferon on virus infection and is activated by 5'-phosphorylated 2'-5'-linked oligoadenylates known as 2-5A [pn5' A2'(p5' A2')mp5'A], resulting in the cleavage of single-stranded RNA predominantly after UpUp and UpAp sequences. To direct 2-5A-dependent RNase to cleave unique RNA sequences, p5' A2' p5' A2'p5'A was covalently linked to an antisense oligonucleotide to yield a chimeric molecule (2-5A:AS). The antisense oligonucleotide component of 2-5A:AS bound a specific RNA sequence while the accompanying 2-5A component activated 2-5A-dependent RNase, thereby causing the cleavage of the RNA in the targeted sequence. This strategy was demonstrated by inducing specific cleavage within a modified human immunodeficiency virus type 1 vif mRNA in a cell-free system from human lymphoblastoid cells. Because 2-5A-dependent RNase is present in most mammalian cells, the control of gene expression based on this technology--including therapies for cancer, viral infections, and certain genetic diseases--can be envisioned.
Torrence
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PF
Section on Biomedical Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
Maitra
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RK
Lesiak
K
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Khamnei
S
S
Zhou
A
A
Silverman
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RH
D000227:Adenine Nucleotides; D017122:Bacteriophage T4 / Q000235:genetics; D001483:Base Sequence; D002678:Chimera; D016341:Genes, vif; D015497:HIV-1 / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense; D009843:Oligoribonucleotides; D010957:Plasmids; D012313:RNA / Q000737:chemistry* / Q000378:metabolism; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism*; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured
D000227:Adenine Nucleotides; D016376:Oligonucleotides, Antisense; D009843:Oligoribonucleotides; D012367:RNA, Viral; C023505:2',5'-oligoadenylate; D012313:RNA; D012260:Ribonucleases