Conformational Propensities of a DNA Hairpin with a Stem Sequence from the c-MYC Promoter
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials
2.2. Circular Dichroism Spectropolarimetry
2.3. UV Spectrophotometry
2.4. Deconvolution of CD Spectra
3. Results
3.1. CD Spectra of GT11C
3.2. CD Spectra of “Pure” Conformations
3.3. Fractions of Individual Conformations
4. Discussion
4.1. Effect of TBA+ Ions on the Duplex-G-Quadruplex Competition
4.2. Thermodynamic Model
4.3. Interpretation of the Temperature Dependences of Fractional Populations
4.4. Distribution of Conformational States at pH 5.0
4.5. Distribution of Conformational States at pH 7.0
4.6. Biological Implications
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
- Lane, A.N.; Chaires, J.B.; Gray, R.D.; Trent, J.O. Stability and kinetics of G-quadruplex structures. Nucleic Acids Res. 2008, 36, 5482–5515. [Google Scholar] [CrossRef] [PubMed]
- Balasubramanian, S.; Hurley, L.H.; Neidle, S. Targeting G-quadruplexes in gene promoters: A novel anticancer strategy? Nat. Rev. Drug Disc. 2011, 10, 261–275. [Google Scholar] [CrossRef] [PubMed]
- Di Antonio, M.; Ponjavic, A.; Radzevicius, A.; Ranasinghe, R.T.; Catalano, M.; Zhang, X.; Shen, J.; Needham, L.M.; Lee, S.F.; Klenerman, D.; et al. Single-molecule visualization of DNA G-quadruplex formation in live cells. Nat. Chem. 2020, 12, 832–837. [Google Scholar] [CrossRef] [PubMed]
- Galli, S.; Melidis, L.; Flynn, S.M.; Varshney, D.; Simeone, A.; Spiegel, J.; Madden, S.K.; Tannahill, D.; Balasubramanian, S. DNA G-quadruplex recognition in vitro and in live cells by a structure-specific nanobody. J. Am. Chem. Soc. 2022, 144, 23096–23103. [Google Scholar] [CrossRef] [PubMed]
- Lam, E.Y.; Beraldi, D.; Tannahill, D.; Balasubramanian, S. G-quadruplex structures are stable and detectable in human genomic DNA. Nat. Commun. 2013, 4, 1796. [Google Scholar] [CrossRef] [PubMed]
- Spiegel, J.; Adhikari, S.; Balasubramanian, S. The structure and function of DNA G-quadruplexes. Trends Chem. 2020, 2, 123–136. [Google Scholar] [CrossRef]
- Varshney, D.; Spiegel, J.; Zyner, K.; Tannahill, D.; Balasubramanian, S. The regulation and functions of DNA and RNA G-quadruplexes. Nat. Rev. Mol. Cell Biol. 2020, 21, 459–474. [Google Scholar] [CrossRef] [PubMed]
- Day, H.A.; Pavlou, P.; Waller, Z.A. i-motif DNA: Structure, stability and targeting with ligands. Bioorg. Med. Chem. 2014, 22, 4407–4418. [Google Scholar] [CrossRef]
- Alba, J.J.; Sadurni, A.; Gargallo, R. Nucleic acid i-motif structures in analytical chemistry. Crit. Rev. Anal. Chem. 2016, 46, 443–454. [Google Scholar] [CrossRef]
- Tateishi-Karimata, H.; Sugimoto, N. Chemical biology of non-canonical structures of nucleic acids for therapeutic applications. Chem. Commun. 2020, 56, 2379–2390. [Google Scholar] [CrossRef]
- Zeraati, M.; Langley, D.B.; Schofield, P.; Moye, A.L.; Rouet, R.; Hughes, W.E.; Bryan, T.M.; Dinger, M.E.; Christ, D. i-motif DNA structures are formed in the nuclei of human cells. Nat. Chem. 2018, 10, 631–637. [Google Scholar] [CrossRef] [PubMed]
- Zanin, I.; Ruggiero, E.; Nicoletto, G.; Lago, S.; Maurizio, I.; Gallina, I.; Richter, S.N. Genome-wide mapping of i-motifs reveals their association with transcription regulation in live human cells. Nucleic Acids Res. 2023, 51, 8309–8321. [Google Scholar] [CrossRef] [PubMed]
- Viskova, P.; Istvankova, E.; Rynes, J.; Dzatko, S.; Loja, T.; Zivkovic, M.L.; Rigo, R.; El-Khoury, R.; Serrano-Chacon, I.; Damha, M.J.; et al. In-cell NMR suggests that DNA i-motif levels are strongly depleted in living human cells. Nat. Commun. 2024, 15, 1992. [Google Scholar] [CrossRef] [PubMed]
- Galli, S.; Flint, G.; Ruzickova, L.; Di Antonio, M. Genome-wide mapping of G-quadruplex DNA: A step-by-step guide to select the most effective method. RSC Chem. Biol. 2024, 5, 426–438. [Google Scholar] [CrossRef] [PubMed]
- Obara, P.; Wolski, P.; Panczyk, T. Insights into the molecular structure, stability, and biological significance of non-canonical DNA forms, with a focus on G-quadruplexes and i-motifs. Molecules 2024, 29, 4683. [Google Scholar] [CrossRef]
- Dell’Oca, M.C.; Quadri, R.; Bernini, G.M.; Menin, L.; Grasso, L.; Rondelli, D.; Yazici, O.; Sertic, S.; Marini, F.; Pellicioli, A.; et al. Spotlight on G-quadruplexes: From structure and modulation to physiological and pathological roles. Int. J. Mol. Sci. 2024, 25, 3162. [Google Scholar] [CrossRef]
- Johnson, F.B. Fundamentals of G-quadruplex biology. Annu. Rep. Med. Chem. 2020, 54, 3–44. [Google Scholar] [PubMed]
- Rhodes, D.; Lipps, H.J. G-quadruplexes and their regulatory roles in biology. Nucleic Acids Res. 2015, 43, 8627–8637. [Google Scholar] [CrossRef]
- Shafer, R.H.; Smirnov, I. Biological aspects of DNA/RNA quadruplexes. Biopolymers 2000, 56, 209–227. [Google Scholar] [CrossRef]
- Hurley, L.H. DNA and its associated processes as targets for cancer therapy. Nat. Rev. Cancer 2002, 2, 188–200. [Google Scholar] [CrossRef]
- Huppert, J.L. Four-stranded DNA: Cancer, gene regulation and drug development. Philos. Trans. R. Soc. A 2007, 365, 2969–2984. [Google Scholar] [CrossRef] [PubMed]
- Huppert, J.L. Four-stranded nucleic acids: Structure, function and targeting of G-quadruplexes. Chem. Soc. Rev. 2008, 37, 1375–1384. [Google Scholar] [CrossRef] [PubMed]
- Oganesian, L.; Bryan, T.M. Physiological relevance of telomeric G-quadruplex formation: A potential drug target. Bioessays 2007, 29, 155–165. [Google Scholar] [CrossRef] [PubMed]
- De Cian, A.; Lacroix, L.; Douarre, C.; Temime-Smaali, N.; Trentesaux, C.; Riou, J.F.; Mergny, J.L. Targeting telomeres and telomerase. Biochimie 2008, 90, 131–155. [Google Scholar] [CrossRef] [PubMed]
- Simone, R.; Fratta, P.; Neidle, S.; Parkinson, G.N.; Isaacs, A.M. G-quadruplexes: Emerging roles in neurodegenerative diseases and the non-coding transcriptome. FEBS Lett. 2015, 589, 1653–1668. [Google Scholar] [CrossRef] [PubMed]
- Millevoi, S.; Moine, H.; Vagner, S. G-quadruplexes in RNA biology. Wiley Interdiscip. Rev. RNA 2012, 3, 495–507. [Google Scholar] [CrossRef] [PubMed]
- Cammas, A.; Dubrac, A.; Morel, B.; Lamaa, A.; Touriol, C.; Teulade-Fichou, M.P.; Prats, H.; Millevoi, S. Stabilization of the G-quadruplex at the VEGF IRES represses cap-independent translation. RNA Biol. 2015, 12, 320–329. [Google Scholar] [CrossRef] [PubMed]
- Shivalingam, A.; Izquierdo, M.A.; Le Marois, A.; Vysniauskas, A.; Suhling, K.; Kuimova, M.K.; Vilar, R. The interactions between a small molecule and G-quadruplexes are visualized by fluorescence lifetime imaging microscopy. Nat. Commun. 2015, 6, 8178. [Google Scholar] [CrossRef]
- Brooks, T.A.; Hurley, L.H. Targeting MYC expression through G-quadruplexes. Genes Cancer 2010, 1, 641–649. [Google Scholar] [CrossRef]
- Collie, G.W.; Parkinson, G.N. The application of DNA and RNA G-quadruplexes to therapeutic medicines. Chem. Soc. Rev. 2011, 40, 5867–5892. [Google Scholar] [CrossRef]
- Bedrat, A.; Lacroix, L.; Mergny, J.L. Re-evaluation of G-quadruplex propensity with G4Hunter. Nucleic Acids Res. 2016, 44, 1746–1759. [Google Scholar] [CrossRef] [PubMed]
- Puig Lombardi, E.; Holmes, A.; Verga, D.; Teulade-Fichou, M.P.; Nicolas, A.; Londono-Vallejo, A. Thermodynamically stable and genetically unstable G-quadruplexes are depleted in genomes across species. Nucleic Acids Res. 2019, 47, 6098–6113. [Google Scholar] [CrossRef] [PubMed]
- Tian, T.; Chen, Y.Q.; Wang, S.R.; Zhou, X. G-quadruplex: A regulator of gene expression and its chemical targeting. Chem 2018, 4, 1314–1344. [Google Scholar] [CrossRef]
- Hansel-Hertsch, R.; Di Antonio, M.; Balasubramanian, S. DNA G-quadruplexes in the human genome: Detection, functions and therapeutic potential. Nat. Rev. Mol. Cell Biol. 2017, 18, 279–284. [Google Scholar] [CrossRef] [PubMed]
- Brooks, T.A.; Kendrick, S.; Hurley, L. Making sense of G-quadruplex and i-motif functions in oncogene promoters. FEBS J. 2010, 277, 3459–3469. [Google Scholar] [CrossRef] [PubMed]
- Völker, J.; Gindikin, V.; Breslauer, K.J. Higher-order DNA secondary structures and their transformations: The hidden complexities of tetrad and quadruplex DNA structures, complexes, and modulatory interactions induced by strand invasion events. Biomolecules 2024, 14, 1532. [Google Scholar] [CrossRef]
- Huppert, J.L.; Balasubramanian, S. G-quadruplexes in promoters throughout the human genome. Nucleic Acids Res. 2007, 35, 406–413. [Google Scholar] [CrossRef] [PubMed]
- Eddy, J.; Maizels, N. Gene function correlates with potential for G4 DNA formation in the human genome. Nucleic Acids Res. 2006, 34, 3887–3896. [Google Scholar] [CrossRef]
- Wright, E.P.; Huppert, J.L.; Waller, Z.A.E. Identification of multiple genomic DNA sequences which form i-motif structures at neutral pH. Nucleic Acids Res. 2017, 45, 2951–2959. [Google Scholar] [CrossRef]
- Dzatko, S.; Krafcikova, M.; Hansel-Hertsch, R.; Fessl, T.; Fiala, R.; Loja, T.; Krafcik, D.; Mergny, J.L.; Foldynova-Trantirkova, S.; Trantirek, L. Evaluation of the stability of DNA i-motifs in the nuclei of living mammalian cells. Angew. Chem. Int. Ed. 2018, 57, 2165–2169. [Google Scholar] [CrossRef]
- Zhou, J.; Wei, C.; Jia, G.; Wang, X.; Feng, Z.; Li, C. Formation of i-motif structure at neutral and slightly alkaline pH. Mol. Biosyst. 2010, 6, 580–586. [Google Scholar] [CrossRef] [PubMed]
- King, J.J.; Irving, K.L.; Evans, C.W.; Chikhale, R.V.; Becker, R.; Morris, C.J.; Pena Martinez, C.D.; Schofield, P.; Christ, D.; Hurley, L.H.; et al. DNA G-quadruplex and i-motif structure formation is interdependent in human cells. J. Am. Chem. Soc. 2020, 142, 20600–20604. [Google Scholar] [CrossRef] [PubMed]
- Chalikian, T.V.; Liu, L.; Macgregor, R.B., Jr. Duplex-tetraplex equilibria in guanine- and cytosine-rich DNA. Biophys. Chem. 2020, 267, 106473. [Google Scholar] [CrossRef] [PubMed]
- Mergny, J.L.; Sen, D. DNA quadruple helices in nanotechnology. Chem. Rev. 2019, 119, 6290–6325. [Google Scholar] [CrossRef] [PubMed]
- Krishnan, Y.; Simmel, F.C. Nucleic acid based molecular devices. Angew. Chem. Int. Ed. 2011, 50, 3124–3156. [Google Scholar] [CrossRef] [PubMed]
- Debnath, M.; Fatma, K.; Dash, J. Chemical regulation of DNA i-motifs for nanobiotechnology and therapeutics. Angew. Chem. Int. Ed. 2019, 58, 2942–2957. [Google Scholar] [CrossRef] [PubMed]
- Liu, L.; Ma, C.; Wells, J.W.; Chalikian, T.V. Conformational preferences of DNA strands from the promoter region of the c-MYC oncogene. J. Phys. Chem. B 2020, 124, 751–762. [Google Scholar] [CrossRef]
- Liu, L.; Zhu, L.; Tong, H.; Su, C.; Wells, J.W.; Chalikian, T.V. Distribution of conformational states adopted by DNA from the promoter regions of the VEGF and Bcl-2 oncogenes. J. Phys. Chem. B 2022, 126, 6654–6670. [Google Scholar] [CrossRef] [PubMed]
- Pandey, A.; Roy, S.; Srivatsan, S.G. Probing the competition between duplex, G-quadruplex and i-motif structures of the oncogenic c-Myc DNA promoter region. Chem. Asian J. 2023, 18, e202300510. [Google Scholar] [CrossRef]
- Ambrus, A.; Chen, D.; Dai, J.X.; Jones, R.A.; Yang, D.Z. Solution structure of the biologically relevant g-quadruplex element in the human c-MYC promoter. implications for G-quadruplex stabilization. Biochemistry 2005, 44, 2048–2058. [Google Scholar] [CrossRef] [PubMed]
- Kim, B.G.; Chalikian, T.V. Thermodynamic linkage analysis of pH-induced folding and unfolding transitions of i-motifs. Biophys. Chem. 2016, 216, 19–22. [Google Scholar] [CrossRef] [PubMed]
- Tataurov, A.V.; You, Y.; Owczarzy, R. Predicting ultraviolet spectrum of single stranded and double stranded deoxyribonucleic acids. Biophys. Chem. 2008, 133, 66–70. [Google Scholar] [CrossRef] [PubMed]
- Rachwal, P.A.; Fox, K.R. Quadruplex melting. Methods 2007, 43, 291–301. [Google Scholar] [CrossRef] [PubMed]
- Mergny, J.L.; Phan, A.T.; Lacroix, L. Following G-quartet formation by UV-spectroscopy. FEBS Lett. 1998, 435, 74–78. [Google Scholar] [CrossRef] [PubMed]
- Benabou, S.; Avino, A.; Eritja, R.; Gonzalez, C.; Gargallo, R. Fundamental aspects of the nucleic acid i-motif structures. RSC Adv. 2014, 4, 26956–26980. [Google Scholar] [CrossRef]
- Dapic, V.; Abdomerovic, V.; Marrington, R.; Peberdy, J.; Rodger, A.; Trent, J.O.; Bates, P.J. Biophysical and biological properties of quadruplex oligodeoxyribonucleotides. Nucleic Acids Res. 2003, 31, 2097–2107. [Google Scholar] [CrossRef] [PubMed]
- Vorlickova, M.; Kejnovska, I.; Bednarova, K.; Renciuk, D.; Kypr, J. Circular dichroism spectroscopy of DNA: From duplexes to quadruplexes. Chirality 2012, 24, 691–698. [Google Scholar] [CrossRef]
- Vorlickova, M.; Kejnovska, I.; Sagi, J.; Renciuk, D.; Bednarova, K.; Motlova, J.; Kypr, J. Circular dichroism and guanine quadruplexes. Methods 2012, 57, 64–75. [Google Scholar] [CrossRef]
- Li, X.; Dubins, D.N.; Volker, J.; Chalikian, T.V. G-quadruplex recognition by tetraalkylammonium ions: A new paradigm for discrimination between parallel and antiparallel G-quadruplexes. J. Phys. Chem. B. 2024, 128, 11144–11150. [Google Scholar] [CrossRef]
- Lipfert, J.; Doniach, S.; Das, R.; Herschlag, D. Understanding nucleic acid-ion interactions. Annu. Rev. Biochem. 2014, 83, 813–841. [Google Scholar] [CrossRef]
- Tateishi-Karimata, H.; Sugimoto, N. Roles of non-canonical structures of nucleic acids in cancer and neurodegenerative diseases. Nucleic Acids Res. 2021, 49, 7839–7855. [Google Scholar] [CrossRef] [PubMed]
- Sugimoto, N.; Endoh, T.; Takahashi, S.; Tateishi-Karimata, H. Chemical biology of double helical and non-double helical nucleic acids: “to B or not to B, that is the question”. Bull. Chem. Soc. Jpn. 2021, 94, 1970–1998. [Google Scholar] [CrossRef]
- Robinson, J.; Raguseo, F.; Nuccio, S.P.; Liano, D.; Di Antonio, M. DNA G-quadruplex structures: More than simple roadblocks to transcription? Nucleic Acids Res. 2021, 49, 8419–8431. [Google Scholar] [CrossRef] [PubMed]
- Armas, P.; David, A.; Calcaterra, N.B. Transcriptional control by G-quadruplexes: In vivo roles and perspectives for specific intervention. Transcription 2017, 8, 21–25. [Google Scholar] [CrossRef] [PubMed]
- Brown, S.L.; Kendrick, S. The i-motif as a molecular target: More than a complementary DNA secondary structure. Pharmaceuticals 2021, 14, 96. [Google Scholar] [CrossRef] [PubMed]
- Esain-Garcia, I.; Kirchner, A.; Melidis, L.; Tavares RC, A.; Dhir, S.; Simeone, A.; Yu, Z.; Madden, S.K.; Hermann, R.; Tannahill, D.; et al. G-quadruplex DNA structure is a positive regulator of MYC transcription. Proc. Natl. Acad. Sci. USA 2024, 121, e2320240121. [Google Scholar] [CrossRef] [PubMed]
- Mendoza, O.; Bourdoncle, A.; Boule, J.B.; Brosh, R.M., Jr.; Mergny, J.L. G-quadruplexes and helicases. Nucleic Acids Res. 2016, 44, 1989–2006. [Google Scholar] [CrossRef]
- Shu, H.; Zhang, R.; Xiao, K.; Yang, J.; Sun, X. G-quadruplex-binding proteins: Promising targets for drug design. Biomolecules 2022, 12, 648. [Google Scholar] [CrossRef]
- Carvalho, J.; Mergny, J.L.; Salgado, G.F.; Queiroz, J.A.; Cruz, C. G-quadruplex, friend or foe: The role of the G-quartet in anticancer strategies. Trends Mol. Med. 2020, 26, 848–861. [Google Scholar] [CrossRef]
pH | ΔHHP | ΔHGQ1 | ΔHGQ2 | ΔHiM |
---|---|---|---|---|
5.0 | 36.7 ± 1.5 | 16.5 ± 1.2 | 12.7 ± 2.3 | 34.9 ± 1.6 |
7.0 | 45.9 ± 2.3 | 35.2 ± 2.2 | ND b | ND c |
[KCl], mM | THP | TGQ1 | TGQ2 b | TiM |
---|---|---|---|---|
0 | 45.8 ± 0.9 | ND c | ND c | 45.9 ± 1.0 |
1 | 50.0 ± 0.9 | 34.7 ± 4.5 d | 0 ± 11.4 d | 49.0 ± 1.1 |
2 | 51.0 ± 0.9 | 50.9 ± 2.3 | 29.5 ± 3.6 | 48.2 ± 1.3 |
5 | 55.2 ± 0.9 | 57.7 ± 2.0 | 28.7 ± 4.3 | 50.1 ± 1.5 |
10 | 61.3 ± 1.0 | 70.9 ± 1.7 | 42.5 ± 5.3 | 53.4 ± 1.9 |
20 | 60.6 ± 1.0 | 68.9 ± 1.7 | 48.0 ± 7.2 | 50.4 ± 2.2 |
50 | 62.4 ± 1.1 | 76.7 ± 1.6 | 58.8 ± 9.7 | 51.5 ± 2.7 |
100 | 67.0 ± 1.0 | 80.0 ± 1.7 | 53.3 ± 10.3 | 50.2 ± 3.0 |
[KCl], mM | THP | TGQ1 | TGQ2 b | TiM c |
---|---|---|---|---|
0 | 58.2 ± 0.8 | ND | ND | ND |
1 | 54.1 ± 1.3 | 68.7 ± 1.1 | ND | ND |
2 | 60.1 ± 1.2 | 73.3 ± 1.2 | ND | ND |
5 | 63.6 ± 1.3 | 81.2 ± 1.2 | ND | ND |
10 | 62.8 ± 1.6 | 85.9 ± 1.2 | ND | ND |
20 | 65.5 ± 1.5 | 87.7 ± 1.2 | ND | ND |
50 | 75.4 ± 1.4 | 93.4 ± 1.4 | ND | ND |
100 | 82.4 ± 1.3 | 92.3 ± 1.4 | ND | ND |
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content. |
© 2025 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Garabet, A.; Prislan, I.; Poklar Ulrih, N.; Wells, J.W.; Chalikian, T.V. Conformational Propensities of a DNA Hairpin with a Stem Sequence from the c-MYC Promoter. Biomolecules 2025, 15, 483. https://doi.org/10.3390/biom15040483
Garabet A, Prislan I, Poklar Ulrih N, Wells JW, Chalikian TV. Conformational Propensities of a DNA Hairpin with a Stem Sequence from the c-MYC Promoter. Biomolecules. 2025; 15(4):483. https://doi.org/10.3390/biom15040483
Chicago/Turabian StyleGarabet, Arees, Iztok Prislan, Nataša Poklar Ulrih, James W. Wells, and Tigran V. Chalikian. 2025. "Conformational Propensities of a DNA Hairpin with a Stem Sequence from the c-MYC Promoter" Biomolecules 15, no. 4: 483. https://doi.org/10.3390/biom15040483
APA StyleGarabet, A., Prislan, I., Poklar Ulrih, N., Wells, J. W., & Chalikian, T. V. (2025). Conformational Propensities of a DNA Hairpin with a Stem Sequence from the c-MYC Promoter. Biomolecules, 15(4), 483. https://doi.org/10.3390/biom15040483