Major G-Quadruplex Form of HIV-1 LTR Reveals a (3 + 1) Folding Topology Containing a Stem-Loop.

Elena Butovskaya,Blaž Bakalar,Anh Tuân Phan,Sara N Richter,Brahim Heddi

doi:10.1021/jacs.8b05332

Elena Butovskaya, Blaž Bakalar + Show 3 more

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https://doi.org/10.1021/jacs.8b05332

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Abstract

Nucleic acids can form noncanonical four-stranded structures called G-quadruplexes. G-quadruplex-forming sequences are found in several genomes including human and viruses. Previous studies showed that the G-rich sequence located in the U3 promoter region of the HIV-1 long terminal repeat (LTR) folds into a set of dynamically interchangeable G-quadruplex structures. G-quadruplexes formed in the LTR could act as silencer elements to regulate viral transcription. Stabilization of LTR G-quadruplexes by G-quadruplex-specific ligands resulted in decreased viral production, suggesting the possibility of targeting viral G-quadruplex structures for antiviral purposes. Among all the G-quadruplexes formed in the LTR sequence, LTR-III was shown to be the major G-quadruplex conformation in vitro. Here we report the NMR structure of LTR-III in K+ solution, revealing the formation of a unique quadruplex–duplex hybrid consisting of a three-layer (3 + 1) G-quadruplex scaffold, a 12-nt diagonal loop containing a conserved duplex-stem, a 3-nt lateral loop, a 1-nt propeller loop, and a V-shaped loop. Our structure showed several distinct features including a quadruplex–duplex junction, representing an attractive motif for drug targeting. The structure solved in this study may be used as a promising target to selectively impair the viral cycle.

Highlights

G-quadruplexes are alternative secondary structures formed by guanine-rich nucleic acids
Using UV, CD, and NMR spectroscopy we investigated the G-quadruplex formation of long terminal repeat (LTR)-III in K+
Our structure of LTR-III G-quadruplex containing a duplex hairpin across a diagonal loop reveals a significant tilting between the helical axis of the duplex and that of the quadruplex, contrasting the feature observed for an artificial hybrid quadruplex−duplex containing a duplex hairpin across a diagonal loop (PDB code 2M91) (Figure S12)

Summary

Introduction

G-quadruplexes are alternative secondary structures formed by guanine-rich nucleic acids. Potential G-quadruplex forming sequences are widespread in the human genome and implicated in key genomic functions, such as transcription, replication, repair, and telomere maintenance.[7−11] overrepresented in the promoter regions of oncogenes, G-quadruplexes act as regulatory elements of gene expression.[9,10] Targeting the G-quadruplex structures in the promoters of oncogenes c-myc, c-kit, and bcl-2 with G-quadruplex-stabilizing agents leads to gene transcription inhibition and decreased levels of gene expression,[12] suggesting G-quadruplexes as promising anticancer targets.[13,14]. In Epstein−Barr virus (EBV) an RNA G-quadruplex regulates translation of EBNA1 mRNA.[16,17]

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