Structural polymorphism within a regulatory element of the human KRAS promoter: formation of G4-DNA recognized by nuclear proteins

Cogoi, Susanna; Paramasivam, Manikandan; Spolaore, Barbara; Xodo, Luigi E.

doi:10.1093/nar/gkn120

Cited by 149 publications

(177 citation statements)

References 67 publications

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“…It occurs by heteromodification of histones, which promotes the decondensation of high order chromatin, and by binding to enhancer/promoter regulatory cis-elements (44). Previous studies have reported that PARP-1 recognizes unusual DNA conformations such as hairpins/cruciforms (47)(48)(49) and the G-quadruplex formed by a cis-element in the human KRAS promoter (8). Interestingly, Ladame and co-workers (50) have shown that PARP-1 undergoes automodification after binding to the c-kit quadruplex.…”

Section: Discussionmentioning

confidence: 99%

The KRAS Promoter Responds to Myc-associated Zinc Finger and Poly(ADP-ribose) Polymerase 1 Proteins, Which Recognize a Critical Quadruplex-forming GA-element

Cogoi

Paramasivam

Membrino

et al. 2010

Journal of Biological Chemistry

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129

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The murine KRAS promoter contains a G-rich nuclease hypersensitive element (GA-element) upstream of the transcription start site that is essential for transcription. Pulldown and chromatin immunoprecipitation assays demonstrate that this GA-element is bound by the Myc-associated zinc finger (MAZ) and poly(ADP-ribose) polymerase 1 (PARP-1) proteins. These proteins are crucial for transcription, because when they are knocked down by short hairpin RNA, transcription is downregulated. This is also the case when the poly(ADP-ribosyl)ation activity of PARP-1 is inhibited by 3,4-dihydro-5-[4-(1-piperidinyl) butoxyl]-1(2H) isoquinolinone. We found that MAZ specifically binds to the duplex and quadruplex conformations of the GA-element, whereas PARP-1 shows specificity only for the G-quadruplex. On the basis of fluorescence resonance energy transfer melting and polymerase stop assays we saw that MAZ stabilizes the KRAS quadruplex. When the capacity of folding in the GA-element is abrogated by specific G 3 T or G 3 A point mutations, KRAS transcription is down-regulated. Conversely, guanidine-modified phthalocyanines, which specifically interact with and stabilize the KRAS G-quadruplex, push the promoter activity up to more than double. Collectively, our data support a transcription mechanism for murine KRAS that involves MAZ, PARP-1 and duplex-quadruplex conformational changes in the promoter GA-element.Guanine-rich sequences have the potential to fold into intramolecular G-quadruplex (or G4-DNA) structures that are stabilized by planar arrays of four guanines paired by Hoogsteen hydrogen bonds (G-tetrad) (1). Quadruplex-forming sequences (QFS) 2 are present in prokaryotic and eukaryotic genomes, promoter regions, micro-and mini-satellite repeats, telomeres, rDNA, and the vertebrate immunoglobulin heavy chain switch regions (2). Recent bioinformatic search analyses have shown a surprisingly high presence in the human genome of QFS, on the order of 3-4 ϫ 10 5 (3, 4). The gene distribution of QFS is highly skewed because tumor suppressor genes have a very low level of QFS, whereas proto-oncogenes have a high level of such sequences (5). There seems to be a correlation between QFS and genomic instability; a low level of QFS in tumor suppressor genes is associated with genomic stability, and a high level is associated with genomic instability (5). Furthermore, the observation that QFS are often located in the region surrounding the transcription start sites of the genes and within cis-elements suggests that they may be involved in transcription regulation. This hypothesis has been formulated for a number of genes including CMYC, KRAS, C-MYB, VEGF, PDGFA, CKIT, and human insulin (6 -13). The best studied G-rich sequence folding into a G-quadruplex, whose function has been correlated with a mechanism of transcription regulation, is the one found in the promoter of the CMYC gene (6). Upstream of the P1 promoter, controlling about 80% of transcription, there is a QFS that can fold into a G-quadruplex. When this quadruplex is d...

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Section: Discussionmentioning

confidence: 99%

The KRAS Promoter Responds to Myc-associated Zinc Finger and Poly(ADP-ribose) Polymerase 1 Proteins, Which Recognize a Critical Quadruplex-forming GA-element

Cogoi

Paramasivam

Membrino

et al. 2010

Journal of Biological Chemistry

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129

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“…As G4-decoy 2998 binds to proteins recognizing G4-proximal (PARP-1, Ku70, hnRNP A1, MAZ)9, it inhibits KRAS transcription by a decoy mechanism. As a consequence, the proliferation and clonogenic potential of Panc-1 cells are reduced by the G4-decoy oligonucleotide12.…”

Section: Resultsmentioning

confidence: 99%

“…Given the strong impact of KRAS G12D on the metabolism of PDAC cells, this oncogene is an attractive target for new anticancer drugs. For a rationale design of anticancer drugs we focused on the KRAS promoter, as it contains three G4 motifs, of which the one most close to TSS, G4-proximal, has been extensively studied8910. G4-proximal is located between −144 and −117 upstream of TSS, overlaps a nuclease hypersensitive element (NHE) and is recognized by several nuclear proteins including MAZ, PARP-1, Ku70/Ku80 and hnRNP A110.…”

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confidence: 99%

“…DMS footprinting and CD experiments showed that sequence G4-proximal folds into a parallel 1/1/11 G-quadruplex with a kinked thymine in one strand, two 1-nt and one 11-nt loops (formed by G-runs 1-2-3-5, Q 1 , T M = 75 °C in 100 mM KCl, Fig. S1)9. Further analyses showed that G4-proximal exhibits a remarkable structural polymorphism, as two truncated G4-proximal sequences composed by G-runs 1-2-3-4 and 2-3-4-5 fold also into a G-quadruplex, respectively with a parallel, T M = 56 °C, 100 mM KCl (Q 2 ) and mixed parallel/antiparallel, T M = 50 °C, 100 mM KCl (Q 3 ) topologies (Fig.…”

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confidence: 99%

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Lipid-modified G4-decoy oligonucleotide anchored to nanoparticles: delivery and bioactivity in pancreatic cancer cells

Cogoi

Jakobsen

Pedersen

et al. 2016

Sci Rep

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KRAS is mutated in >90% of pancreatic ductal adenocarcinomas. As its inactivation leads to tumour regression, mutant KRAS is considered an attractive target for anticancer drugs. In this study we report a new delivery strategy for a G4-decoy oligonucleotide that sequesters MAZ, a transcription factor essential for KRAS transcription. It is based on the use of palmitoyl-oleyl-phosphatidylcholine (POPC) liposomes functionalized with lipid-modified G4-decoy oligonucleotides and a lipid-modified cell penetrating TAT peptide. The potency of the strategy in pancreatic cancer cells is demonstrated by cell cytometry, confocal microscopy, clonogenic and qRT-PCR assays.

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“…We focused on DNA quadruplex sequences and used the human telomeric G-quadruplex (H-telo) [6] and the sequences of the promoter quadruplexes of a selection of genes, namely c-kit (which contains the two G-quadruplex sequences c-kit1 [39] and c-kit2 [40] ), c-myc , [5b, 41] bcl2 [42] and k-ras . [43] These different sequences differ in composition, in the number of nucleotides involved in each loop, and adopt different quadruplex conformations, potentially allowing differential recognition by small molecules that do not target the tetrads per se . As a control we used a sequence that forms a duplex DNA in solution.…”

Section: Resultsmentioning

confidence: 99%

Targeting DNA G‐Quadruplexes with Helical Small Molecules

et al. 2014

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We previously identified quinoline-based oligoamide helical foldamers and a trimeric macrocycle as selective ligands of DNA quadruplexes. Their helical structure may permit the targeting of the backbone loops and grooves of G-quadruplexes instead of the G-tetrads. Given the vast array of morphologies G-quadruplex structures can adopt, this may be a way to elicit sequence selective binding. Herein, we describe the design and synthesis of molecules based on macrocyclic and helically folded oligoamides. We tested their ability to interact with the human telomeric G-quadruplex and an array of promoter G-quadruplexes using Förster Resonance Energy Transfer (FRET) melting assay and single molecule FRET. Our results show that they constitute very potent ligands, comparable to the best reported in the literature. Their mode of interaction differs from that of traditional tetrad binders, opening avenues for the development of molecules specific for certain G-quadruplex conformations.

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Structural polymorphism within a regulatory element of the human KRAS promoter: formation of G4-DNA recognized by nuclear proteins

Cited by 149 publications

References 67 publications

The KRAS Promoter Responds to Myc-associated Zinc Finger and Poly(ADP-ribose) Polymerase 1 Proteins, Which Recognize a Critical Quadruplex-forming GA-element

The KRAS Promoter Responds to Myc-associated Zinc Finger and Poly(ADP-ribose) Polymerase 1 Proteins, Which Recognize a Critical Quadruplex-forming GA-element

Lipid-modified G4-decoy oligonucleotide anchored to nanoparticles: delivery and bioactivity in pancreatic cancer cells

Targeting DNA G‐Quadruplexes with Helical Small Molecules

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