U3 Region in the HIV‑1 Genome Adopts a G‑Quadruplex
Structure in Its RNA and DNA Sequence
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Abstract
Genomic regions rich in G residues
are prone to adopt G-quadruplex
structure. Multiple Sp1-binding motifs arranged in tandem have been
suggested to form this structure in promoters of cancer-related genes.
Here, we demonstrate that the G-rich proviral DNA sequence of the
HIV-1 U3 region, which serves as a promoter of viral transcription,
adopts a G-quadruplex structure. The sequence contains three binding
elements for transcription factor Sp1, which is involved in the regulation
of HIV-1 latency, reactivation, and high-level virus expression. We
show that the three Sp1 binding motifs can adopt different forms of
G-quadruplex structure and that the Sp1 protein can recognize and
bind to its site folded into a G-quadruplex. In addition, a c-kit2
specific antibody, designated hf2, binds to two different G-quadruplexes
formed in Sp1 sites. Since U3 is encoded at both viral genomic ends,
the G-rich sequence is also present in the RNA genome. We demonstrate
that the RNA sequence of U3 forms dimers with characteristics known
for intermolecular G-quadruplexes. Together with previous reports
showing G-quadruplex dimers in the <i>gag</i> and cPPT regions,
these results suggest that integrity of the two viral genomes is maintained
through numerous intermolecular G-quadruplexes formed in different
RNA genome locations. Reconstituted reverse transcription shows that
the potassium-dependent structure formed in U3 RNA facilitates RT
template switching, suggesting that the G-quadruplex contributes to
recombination in U3