RNASE-H IS RESPONSIBLE FOR THE NONSPECIFIC INHIBITION OF IN-VITRO TRANSLATION BY 2'-O-ALKYL CHIMERIC OLIGONUCLEOTIDES - HIGH-AFFINITY OR SELECTIVITY, A DILEMMA TO DESIGN ANTISENSE OLIGOMERS

被引:41
作者
LARROUY, B
BOIZIAU, C
SPROAT, B
TOULME, JJ
机构
[1] UNIV BORDEAUX 2,INSERM,U386,LAB MOLEC BIOPHYS,F-33076 BORDEAUX,FRANCE
[2] EUROPEAN MOLEC BIOL LAB,D-69117 HEIDELBERG,GERMANY
关键词
D O I
10.1093/nar/23.17.3434
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Ribonuclease H (RNase H) which recognizes and cleaves the RNA strand of mismatched RNA-DNA heteroduplexes can induce non-specific effects of antisense oligonucleotides, In a previous paper [Larrouy et al, (1992), Gene, 121, 189-194], we demonstrated that ODN1, a phosphodiester 15mer targeted to the AUG initiation region of alpha-globin mRNA, inhibited non-specifically P-globin synthesis in wheat germ extract due to RNase H-mediated cleavage of beta-globin mRNA, Specificity was restored by using MP-ODN2, a methylphosphonate-phosphodiester sandwich analogue of ODN1, which limited RNase H activity on non-perfect hybrids, We report here that 2'-O-alkyl RNA-phosphodiester DNA sandwich analogues of ODN1, with the same phosphodiester window as MP-ODN2, are non-specific inhibitors of globin synthesis in wheat germ extract, whatever the substituent (methyl, allyl or butyl) on the 2'-OH, These sandwich oligomers induced the cleavage of non-target beta-globin RNA sites, similarly to the unmodified parent oligomer ODN1, This is likely due to the increased affinity of 2'-O-alkyl-ODN2 chimeric oligomers for both fully and partly complementary RNA, compared to MP-ODN2, In contrast, the fully modified 2'-O-methyl analogue of ODN1 was a very effective and highly specific antisense sequence, This was ascribed to its inability (i) to induce RNA cleavage by RNase H and (ii) to physically prevent the elongation of the polypeptide chain.
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页码:3434 / 3440
页数:7
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