Via Research Recognition Day 2024 VCOM-Carolinas

Biomedical Studies

^dZh dhZ > zE D/ ^ K& KE K' E WZKDKd ZͲWZKy/D > E ͗ 'ͲYh ZhW> y ^ E /ͲDKd/&^ /E E Z Z 'h> d/KE Rachel Daley, OMS-II 1 , Rebecca Corallo, OMS-II, Krishna Patel, OMS-II, Sundeep Bhanot, OMS-I, Shane Donahue, OMS-I, Lauren Hiers, OMS-I, Daniel Ross, Olivia Lewis, Bidyut K. Mohanty, PhD. Edward Via College of Osteopathic Medicine, Spartanburg, SC. . Abstract Results Results Continued

Background: The aberrant expression of oncogenes is a well-recognized hallmark in cancer pathogenesis. A pivotal area of focus in cancer research lies in understanding the role of oncogenes, particularly within their promoter-proximal regions (Figure 1A). These regions contain polyguanine/polycytosine-rich DNA sequences, which conventionally form Watson-Crick double-stranded base pairs. Yet, they also exhibit the potential to adopt noncanonical structures, such as G-quadruplexes (G4s) and intercalating motifs (i-Motifs) 1 (Figure 1C). These unconventional configurations have the potential to disrupt genome stability and exert influence on gene regulation, thereby permitting uncontrolled cell growth – the basis of numerous cancers.

Figure 5. 12% Non-denaturing polyacrylamide gel electrophoresis analysis of various DNA samples.

Conclusion: In conclusion, our study demonstrates that polycytosine polyguanine-rich DNA sequences exhibit a pH-dependent preference for i-motif and G-quadruplex structures, respectively. Also, the presence of polyguanine-rich DNA at pH 5.5 has a significant impact on i-motif formation. These findings suggest a preference for the environment-dependent formation of i-Motifs and G-quadruplexes, offering valuable insights into the structural dynamics of the promoter proximal regions of these various oncogenes. Future directions: Understanding the behavior of these regions will be imperative to determine their potential implication in regulating oncogenes. The findings of this study will lay the groundwork for further investigations into the structural dynamics of these DNA sequences. These data will assist in identifying specific targets for therapeutic intervention in oncology. rich DNA at pH 5.5 led to a reduction in i-motif formation by the polycytosine-rich DNA, indicating a dynamic interplay between these structures. Conclusions Results: CD analysis of polycytosine-rich DNA sequences of BCL-2, EGFR, HIF1- α, PDGF, and VEGF unveil distinct i-Motif patterns (Figure 3). Additionally, the polyguanine-rich sequence of BCL-2 displayed a unique G-quadruplex pattern (Figure 4) and its effect on i motif. At pH 5.5, all cytosine-rich DNA sequences formed i-motifs, while guanine-rich DNA showed minimal G-quadruplex formation. However, at pH 7.5, the guanine-rich DNA sequences formed distinct G4 structures. Interestingly, increasing the concentrations of guanine

Figure 1. G4- and iM-forming sequences at oncogene promoters. A. Promoter Proximal Region of oncogenes, B. Proximal Promoter Sequences of Bcl-2, EGFR, HIC, PDGF, and VEGF, C. G-quadruplex and i-Motif Structures

Figure 3. CD analysis of polycytosine-rich DNAs of PDGF, EFGR, VEGF, and HIF1- α reveals distinct i-motif patterns͘

Objective and Methods

Objective: This study aims to investigate the dynamics of G4s and i-Motifs in the promoter-proximal regions of various oncogenes, including Bcl-2, EGFR, HIF1- α, PDGF, and VEGF. We employ Circular Dichroism (CD) spectroscopy (Figure 2A) along with other biochemical and biophysical techniques to explore how environmental conditions influence the formation of G4s and i-Motifs. The CD spectroscopy performed on the promoter proximal regions of these oncogenes reveals distinct patterns of G4s and i Motifs. Our study also explores the effects of various factors, including pH conditions, the complementary DNA strand, and protein factors on the formation of these structures. 2A . 2B .

Figure 2. Circular dichroism spectroscopy. A. Jasco J-1500 CD Spectroscope, B. CD Spectroscope Principle Methods: We used CD spectroscopy to monitor DNA secondary structures in the promoter-proximal region of various oncogenes under differing environmental conditions. G4s and i-motifs are noncanonical DNA structures that display characteristic CD spectra that differ from DNA duplex structures, and from one another. 2

References and Acknowledgements

Figure 4. CD analysis showing the effects of increasing concentrations of polyguanine-rich BCL2 DNA on i-motif formed by polycytosine-rich BCL2 DNA.

BKM is funded by VCOM’s 2024 REAP grant (#1032453).

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2024 Research Recognition Day