Extension of DNA triple helix formation to a neighbouring (AT)<sub><i>n</i></sub> site

Chandler, Simon; Fox, Keith R.

doi:10.1016/0014-5793(95)00069-l

Cited by 14 publications

(14 citation statements)

References 23 publications

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“…Furthermore, our study shows that the stability of all base triplets is within four-fold of T•A–T (Figure 1H and Table 1), whereas quantitative affinity cleavage assays showed all non-canonical base triplets as 15- to 1500-fold less stable than T•A–T (24). The G•T–A base triplet is typically one of the most stable non-canonical D•D–D base triplets (24,26,40,48–50); however, our study showed G•T–A as being comparable to most non-canonical base triplets (Figure 1H and Table 1). The similar relative stability of our D•D–D base triplets is likely because our D•D–D triple helix is too stable for our binding assay to detect stability changes for only a single base triplet.…”

Section: Discussioncontrasting

confidence: 65%

“…molecularity, length of the triple helix, location of varying Z•X–Y base triplet, sequence composition, ratio of canonical T•A–T:C•G–C base triplets and distribution of pH-sensitive C•G–C base triplets), conditions of binding reaction (i.e. buffer species and pH, salt identity and concentrations, polyamines, solvent, molecular crowding agents, temperature and equilibration time) and type of assay used to measure relative stabilities (24–26,40,48–50). Similar study-to-study variations have been observed for the stability of D•D–D base triplets and base pairs in dsRNA, both of which vary greatly depending on neighboring base triplets or base pairs (46,51,52).…”

Section: Discussionmentioning

confidence: 99%

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Stability of an RNA•DNA–DNA triple helix depends on base triplet composition and length of the RNA third strand

Kunkler

Hulewicz

Hickman

et al. 2019

Nucleic Acids Research

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Recent studies suggest noncoding RNAs interact with genomic DNA, forming an RNA•DNA–DNA triple helix that regulates gene expression. However, base triplet composition of pyrimidine motif RNA•DNA–DNA triple helices is not well understood beyond the canonical U•A–T and C•G–C base triplets. Using native gel-shift assays, the relative stability of 16 different base triplets at a single position, Z•X–Y (where Z = C, U, A, G and X–Y = A–T, G–C, T–A, C–G), in an RNA•DNA–DNA triple helix was determined. The canonical U•A–T and C•G–C base triplets were the most stable, while three non-canonical base triplets completely disrupted triple-helix formation. We further show that our RNA•DNA–DNA triple helix can tolerate up to two consecutive non-canonical A•G–C base triplets. Additionally, the RNA third strand must be at least 19 nucleotides to form an RNA•DNA–DNA triple helix but increasing the length to 27 nucleotides does not increase stability. The relative stability of 16 different base triplets in DNA•DNA–DNA and RNA•RNA–RNA triple helices was distinctly different from those in RNA•DNA–DNA triple helices, showing that base triplet stability depends on strand composition being DNA and/or RNA. Multiple factors influence the stability of triple helices, emphasizing the importance of experimentally validating formation of computationally predicted triple helices.

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Section: Discussioncontrasting

confidence: 65%

Section: Discussionmentioning

confidence: 99%

Stability of an RNA•DNA–DNA triple helix depends on base triplet composition and length of the RNA third strand

Kunkler

Hulewicz

Hickman

et al. 2019

Nucleic Acids Research

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“…absence of BePI [126]. Duplex regions of (AT) n have also been targeted with GT-containing oligonucleotides in the presence of the naphthylquinoline triplex-binding ligand, generating complexes containing alternating G q TA and T q AT triplets [127]. Although blocks of alternating T q AT and G q TA triplets alone are not stable, even in the presence a triplex-binding ligand, these complexes can be stabilized by attaching this region to a block of consecutive T q AT triplets, to which the ligand is thought to bind.…”

Section: Triplex Binding Ligandsmentioning

confidence: 99%

Targeting DNA with Triplexes

Fox¹

2000

CMC

191

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The formation of intermolecular DNA triple helices offers the possibility of designing compounds with extensive sequence recognition properties which may be useful as antigene agents or tools in molecular biology. In these structures a third strand oligonucleotide binds in the DNA major groove, making specific contacts with substituents on the exposed faces of the base pairs. Although triplexes form with exquisite specificity their use suffers from several drawbacks. Two limitations of this approach, which are considered in this review are, firstly that conditions of low pH are necessary for formation of the C+*GC triplet, and secondly that these structures are often less stable than their duplex counterparts. This review outlines the strategies that have been employed to overcome these drawbacks. The pH problem is addressed by considering the various DNA base analogues that have been used to recognise GC base pairs in a pH independent fashion, and discusses the benefits and limitations of each analogue. Triplex stability can be increased by using novel base analogues, backbone modifications and the use of triplex-specific binding ligands.

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“…Formation of triplexes with mixed GT-TFMs have been described to form stable triplexes in acidic and neutral pH 37,38 . The W-TFO library (A/T) exhibits stretches of adenines as well as thymines (pvalue ≤ 4.9x10 -324 ).…”

Section: Triplex-seq Reveals New Triplex Forming Motifs (Tfms)mentioning

confidence: 99%

Identifying triplex binding rulesin vitroleads to creation of a new synthetic regulatory toolin vivo

Kaufmann

Willinger

Eden

et al. 2019

Preprint

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Nature provides a rich toolbox of dynamic nucleic acid structures that are widespread in cells and affect multiple biological processes 1 . Recently, non-canonical structures gained renewed scientific and biotechnological interest 2,3 . One particularly intriguing form of such structures are triplexes 4 in which a single-stranded nucleic acid molecule interacts via Hoogsteen bonds with a DNA/RNA double helix 5 . Despite extensive research in vitro 6-9 , the underlying rules for triplex formation remain debated and evidence for triplexes in vivo is circumstantial 10-12 . Here, we demonstrate the development of a deep-sequencing platform termed Triplex-Seq to systematically refine the DNA triplex code and identify high affinity triplex forming oligo (TFO)variants. We identified a preference for short G-rich motifs using an oligo-library with a mix of all four bases. These high-information content motifs formed specific high-affinity triplexes in a pH-independent manner and stability was increased with G-rich double-stranded molecules. We then conjugated one high-affinity and one low-affinity variant to a VP48 peptide and studied these synthetic biomolecules in mammalian cells. Using these peptide-oligo constructs (POCs), we demonstrated possible triplex-induced down-regulation activity in 544 differentially expressed genes. Our results show that deep-sequencing platforms can substantially expand our understanding of triplex binding rules, which in turn has led to the development of a functional non-genetically encoded regulatory tool for in vivo applications.

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Extension of DNA triple helix formation to a neighbouring (AT)_n site

Cited by 14 publications

References 23 publications

Stability of an RNA•DNA–DNA triple helix depends on base triplet composition and length of the RNA third strand

Stability of an RNA•DNA–DNA triple helix depends on base triplet composition and length of the RNA third strand

Targeting DNA with Triplexes

Identifying triplex binding rulesin vitroleads to creation of a new synthetic regulatory toolin vivo

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Extension of DNA triple helix formation to a neighbouring (AT)n site

Cited by 14 publications

References 23 publications

Stability of an RNA•DNA–DNA triple helix depends on base triplet composition and length of the RNA third strand

Stability of an RNA•DNA–DNA triple helix depends on base triplet composition and length of the RNA third strand

Targeting DNA with Triplexes

Identifying triplex binding rulesin vitroleads to creation of a new synthetic regulatory toolin vivo

Contact Info

Product

Resources

About

Extension of DNA triple helix formation to a neighbouring (AT)_n site