A SMALL CIRCULAR TAR RNA DECOY SPECIFICALLY INHIBITS TAT-ACTIVATED HIV-1 TRANSCRIPTION

Linear TAR RNA has previously been used as a decoy to inhibit HIV-1 tr anscription in vitro and HIV-1 replication in vivo, A 48 nucleotide ci rcular RNA containing the stem, bulge and loop of the HIV-1 TAR elemen t was synthesized using the self-splicing activity of a group I permut ed intron-exon and was tested for its ability to function as a TAR dec oy in vitro. This small circular TAR molecule was exceptionally stable in HeLa nuclear extracts, whereas a similar linear TAR molecule was r apidly degraded. The TAR circle bound specifically to Tfr38, a peptide containing the TAR-binding region of Tat. The ability of Tat to trans -activate transcription from the HIV-1 promoter in vitro was efficient ly inhibited by circular TAR RNA but not by TAR circles that contained either bulge or loop mutations, TAR circles did not inhibit trans-act ivation exclusively by binding to Tat since this inhibition was not re versed by adding excess Tat to the transcription reaction. Together, t hese data suggest that TAR circles act as decoys that inhibit trans-ac tivation by binding to Tat and at least one cellular factor, These dat a also demonstrate the utility of small circular RNA molecules as tool s for biochemical studies.