OLIGONUCLEOTIDE ANALOGUE INTERFERENCE WITH THE HIV-1 TAT PROTEIN-TAR RNA INTERACTION

Abstract

The HIV-1 Tat protein interaction with its RNA recognition sequence TAR is an important drug target and model system for the development of specific RNA-protein inhibitors. 2′-O-methyl oligoribonucleotides complementary to the TAR apical stem-loop effectively block Tat binding in vitro. Substitution by 5-propynylC or 5-methylC LNA monomeric units into a 12-mer 2′-O-methyl oligoribonucleotide leads to stronger inhibition, as does a 12-mer PNA. 10–16 mer 2′-O-methyl oligoribonucleotides give sequence- and dose-dependent inhibition of Tat-dependent transcription of an HIV DNA template in HeLa cell nuclear extract. Inhibition is maintained for the substituted 12-mer analogues but is poorer for PNA and is not correlated with TAR binding strength.

ACKNOWLEDGMENTS

We thank Tony Lowe for HeLa cell nuclear extract, Jon Karn for Tat protein and Mark Churcher and Tom Barlow for advice on in vitro transcription. Ms Britta M. Dahl is thanked for LNA synthesis. VKR and JW thank the Danish Natural Science Research Council and the Danish Technical Research Council for financial support.