The proximal promoter region of the human vascular endothelial growth factor gene has a G-quadruplex structure that can be targeted by G-quadruplex–interactive agents

Abstract

Previous studies on the functional analysis of the human vascular endothelial growth factor (VEGF) promoter using the full-length VEGF promoter reporter revealed that the proximal 36-bp region (À85 to À50 relative to transcription initiation site) is essential for basal or inducible VEGF promoter activity in several human cancer cells. This region consists of a polypurine (guanine) tract that contains four runs of at least three contiguous guanines separated by one or more bases, thus conforming to a general motif capable of forming an intramolecular Gquadruplex. Here, we show that the G-rich strand in this region is able to form an intramolecular propeller-type parallel-stranded G-quadruplex structure in vitro by using the electrophoretic mobility shift assay, dimethyl sulfate footprinting technique, the DNA polymerase stop assay, circular dichroism spectroscopy, and computer-aided molecular modeling. Two well-known G-quadruplexinteractive agents, TMPyP4 and Se2SAP, stabilize Gquadruplex structures formed by this sequence in the presence of a potassium ion, although Se2SAP is at least 10-fold more effective in binding to the G-quadruplex than TMPyP4. Between these two agents, Se2SAP better suppresses VEGF transcription in different cancer cell lines, including HEC1A and MDA-MB-231. Collectively, our results provide evidence that specific G-quadruplex structures can be formed in the VEGF promoter region, and that the transcription of this gene can be controlled by ligand-mediated G-quadruplex stabilization. Our results also provide further support for the idea that G-quadruplex structures may play structural roles in vivo and therefore might provide insight into novel methodologies for rational drug design. [Mol Cancer Ther 2008;7(4):880 -9]

Cellular uptake and binding of guanidine-modified phthalocyanines to KRAS/HRASG-quadruplexes

“…GPcs also exhibited good cellular uptake into living cells and suppressed luciferase expression. These results are consistent with G-quadruplex-mediated promoter inhibition, [4][5][6][7][8][9] and provide motivation to further explore the anticancer potential of GPcs.…”

Oxidative DNA damage is epigenetic by regulating gene transcription via base excision repair

“…Previous experiments found the VEGF PQS to be bound by three equivalents of the SP1 transcription factor, and G4 formation is also involved in activation of this gene (8,16). Decoupling of G-quadruplex formation from SP1 binding is complicated by the observation that this transcription factor binds both duplex and G-quadruplex folded DNA of the same sequence with similar affinity (17); additionally, OG in duplex DNA decreases SP1 binding by 70-90% depending on the position (2), and the impact in G-quadruplex DNA is not known.…”

“…Another notable example appears when rat pulmonary artery endothelial cells are subject to hypoxia; a strong positive correlation between OG in promoter regions and elevated expression of >100 genes was observed (7). One gene in particular is vascular endothelial growth factor (VEGF), for which OG was found in the G-rich potential G-quadruplex sequence (PQS) (7) demonstrated to be responsible for transcriptional regulation of the gene (8). Strong cellular evidence for enhancement of transcription via folding of promoter G-quadruplexes was recently demonstrated by the Balasubramanian laboratory (9).…”

Assessment of gene promoter G-quadruplex binding and modulation by a naphthalene diimide derivative in tumor cells

“…Indeed, G-4s can be induced and stabilized by small molecules generally presenting large flat aromatic moieties and cationic side chains. Treatment of cells with G-4-stabilizing compounds has been shown to alter expression of many genes harbouring such sequences (5,9,(13)(14)(15)(16). To date, a diverse array of G-4 stabilizing compounds has been identified (17,18).…”