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Abstract Human telomeres are composed of TTAGGG repeats that can fold into G-quadruplexes (G4s). G4s can form several different conformations, including parallel, antiparallel 2 + 2 chair, antiparallel 2 + 2 basket, and 3 + 1 parallel/antiparallel. Telomeres are composed of deoxyribonucleotide monophosphates; however, telomerase has been shown to insert ribonucleotide monophosphates (rNMPs) as efficiently as replicative DNA polymerases. Non-telomeric rNMP insertions are deleterious, but the effect on telomeres remains under explored. We systematically investigated 16 variants of the G4-forming telomeric sequence (TTAGGG)4 containing a single rNMP substitution. We generally found that rNMP substitution of the first dG in a repeat (TTAGGG)4 altered the G4 conformation. Incorporation of a rNMP also perturbed G4 folding dynamics, decreasing the population of stably folded molecules and promoting rapid structural transitions. Depending on the rNMP position, we further observed a reduction in overall thermal stability. Additionally, RNase H2, the initiator of ribonucleotide excision repair, had reduced cleavage of rNMPs in G4s, and could only cleave rNMPs at more accessible positions within the G4. Cumulatively, we show that the insertion of rNMPs in telomeric sequences alters the conformation and stability of G4s. This could lead to deleterious effects on telomeric integrity, and these changes may persist due to the difficulty of repairing rNMPs within G4s.
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Long G4-rich enhancers target promoters via a G4 DNA-based mechanism
Abstract Several studies have now described instances where G-rich sequences in promoters and enhancers regulate gene expression through forming G-quadruplex (G4) structures. Relatedly, our group recently identified 301 long genomic stretches significantly enriched for minimal G4 motifs (LG4s) in humans and found the majority of these overlap annotated enhancers, and furthermore, that the promoters regulated by these LG4 enhancers are similarly enriched with G4-capable sequences. While the generally accepted model for enhancer:promoter specificity maintains that interactions are dictated by enhancer- and promoter-bound transcriptional activator proteins, the current study tested an alternative hypothesis: that LG4 enhancers interact with cognate promoters via a direct G4:G4 DNA-based mechanism. This work establishes the nuclear proximity of LG4 enhancer:promoter pairs, biochemically demonstrates the ability of individual LG4 single-stranded DNAs (ssDNAs) to directly interact target promoter ssDNAs, and confirms that these interactions, as well as the ability of LG4 enhancers to activate target promoters in culture, are mediated by G4 DNA.
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- Award ID(s):
- 2223547
- PAR ID:
- 10613990
- Author(s) / Creator(s):
- ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; more »
- Publisher / Repository:
- Oxford University Press
- Date Published:
- Journal Name:
- Nucleic Acids Research
- Volume:
- 53
- Issue:
- 2
- ISSN:
- 0305-1048
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
-
Accepted Manuscript
- Journal Article:
- https://doi.org/10.1093/nar/gkae1180
