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Figure 1 | Retrovirology

Figure 1

From: Opening of the TAR hairpin in the HIV-1 genome causes aberrant RNA dimerization and packaging

Figure 1

The HIV-rtTA genome and mutations in the TAR hairpin. (A) The HIV-rtTA proviral DNA genome and the organization and secondary structure of the leader RNA are shown. In HIV-rtTA, the Tat-TAR axis of transcription regulation was inactivated by mutation of both Tat and TAR (tatm and TARm; crossed boxes). Transcription and replication of the virus were made dox-dependent by the introduction of tetO elements in the U3 promoter region and replacing the Nef gene by the rtTA gene. All HIV-rtTA DNA constructs used in this study contained an SV40-derived polyadenylation signal immediately downstream of the viral genome in the plasmid backbone [17.]. The untranslated leader RNA of HIV-1 (+1/+368) can fold several stem-loop structures with important regulatory functions (TAR; polyA: polyadenylation signal; PAS: primer activation signal; PBS: primer binding site; DIS: dimerization initiation signal; SD: splice donor; Ψ: RNA packaging signal; AUG: translation start codon of gag; see text for details). (B) The TARm hairpin with bulge and loop mutations (in red) as present in HIV-rtTA is shown on the left side. The TARm sequence is partially deleted in the mutants A (deletion of nucleotides 11–24 [Δ11-24]), B (Δ39-48) and the double mutant AB (Δ11-24 + Δ39-48; deletions indicated by a grey box). The structure (at 37°C) and thermodynamic stability (ΔG in kcal/mole) of the TAR hairpins as predicted by the RNA Mfold program version 3.5 [18] are shown. Previous in vitro RNA-structure probing of leader transcripts indicated that the 3’ terminal GGGAACC nucleotides of the A and B mutated TAR elements (but not of the TARm and AB variants) interact with nucleotides immediately downstream of the polyA hairpin, which results in further destabilization of the TAR element [19.]

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