Spying HIV-1 G-quadruplexes with nucleoside probes: a unique hotspot for antiviral therapy
Work done in the lab of Prof. Seergazhi G. Srivatsan at the Indian Institute of Science Education and Research, Pune
Sarupa Roy has completed her Bachelor’s (Chemistry Hons.) from Jadavpur University and received her Master’s degree at the Indian Institute of Technology, Guwahati. She is a senior graduate student working under the supervision of Prof. S. G. Srivatsan at the Indian Institute of Science Education and Research, India, Pune. Her research interests revolve around probing viral secondary structures with nucleoside analogs.
Author Interview
How would you explain your research outcomes to the non-scientific community?
Whenever we think of nucleic acids, the first structure that pops into our head is the double-stranded DNA, but it is way beyond that. They can form a plethora of other secondary structures. One such secondary structure is G-quadruplexes (GQs), formed by the stretches of guanines in the DNA or RNA. Four guanines interact via special Hoogsteen H-bonds in the presence of metal ions such as potassium and sodium to form a tetrad. When two or more tetrads stack, they form these GQ structures. Like speed bumpers which slow down the pace of vehicles, GQs regulate the various cellular checkpoints such as transcription, translation, and gene regulation, modulating the speed of various polymerases. Recently, they have been found to be present in viral genomes as well which resulted in a spike of interest in G-quadruplexes and viruses.
As we are aware of the fatal disease-causing HIV-1 virus despite using a cocktail of drugs, remains a major challenge. Henceforth, understanding the conserved regulatory regions of the viral genome would surely assist in developing novel therapeutics. In this context, we have tried to comprehensively understand the secondary structures formed by the long terminal repeat (LTR) present in the promoter region of the HIV-1 provirus. We developed a smart probe system obtained by conjugating the fluorobenzofuran ring to the thymidine base which acts as a dual-purpose GQ sensor, employing both fluorescence and NMR. Whereas a hairpin sensor is obtained by integrating a fluorine label in the thymidine residue. These are highly environment-sensitive, meaning can report the changes in the local environment. These probes, when put in the loop region of the structures, behaved like CCTV cameras and assisted in interrogating the secondary structures with fluorescence and NMR (Figure 1 (a). Interestingly, with the powerful fluorine label, we evaluated the hybrid-type folding consisting of hairpin juxtaposed to GQ (Figure 1 (b)) acquired inside the cellular environment. With the responsiveness of the probe and polymerase stop assay, we also mapped the druggable space and the effect of small molecules on viral machinery (Figure 1 (c)). With that, we propose a bimodal ligand tethered by a linker that could selectively target these unique caricatures and stop the viral replication.
Our probe system not only allows the study of the viral G-quadruplexes but has also generated a screening platform to detect binders interacting with different domains of the structures.
“This finding paves the way to propose a model for developing specific binders that can effectively stabilize these structures and control viral proliferation. “
How do these findings contribute to your research area?
Around the globe, drugs to cure HIV-1 are being developed, but none can effectively eradicate the virus. Hence, there is an urgent need to develop specific binders. Our comprehensive study has helped deconvolute the complex structures, demonstrating the existence of hairpin-attached G-quadruplexes (GQs) in the cellular environment. This finding paves the way to propose a model for developing specific binders that can effectively stabilize these structures and control viral proliferation.
What was the exciting moment during your research?
Nonetheless, an array of binders are selective to G-quadruplexes (GQs) and can stabilize these structures. Interestingly, utilizing a sensitive probe system makes it possible to monitor the interaction of hairpin/G-tetrad selective binders with a high degree of precision. This technique allows for the interactions to be observed not only qualitatively, providing visual confirmation of binding events, but also quantitatively, offering detailed measurements of binding affinity and dynamics. This dual capability significantly enhances our ability to study and understand the binding mechanisms, ultimately aiding in the development of more effective therapeutic agents targeting GQs.
What do you hope to do next?
As the landscape of the viral genome is wide, we next want to explore and understand the various dynamic structures present in RNA viruses with responsive nucleoside analogs that will placidly detect different dynamic structures present in the conserved region.
Where do you seek scientific inspiration from?
I seek scientific inspiration from two pivotal figures in my life. First, my high school chemistry teacher who has always encouraged me to be curious and explore the wonders of science. His enthusiasm and support sparked my initial interest in chemistry and instilled in me a love for scientific discovery. Second, my PhD mentor, whose passion and dedication to scientific endeavors have been truly inspiring. His unwavering commitment to research and his guidance has profoundly shaped my approach to science and fueled my ambition to contribute meaningfully to the field.
How do you intend to help Indian science improve?
India is rapidly emerging as a significant player in the realm of science, showcasing remarkable advancements and innovations. Leveraging the collaborative spirit and expertise available within this dynamic environment, I aspire to develop comprehensive studies on viruses. By collaborating with fellow scientists and researchers, I aim to design and synthesize small molecule binders that can efficiently target and neutralize viral threats. This endeavor not only holds the potential to contribute to global health but also positions India at the forefront of cutting-edge virological research and therapeutic development.
Reference: Roy, S.; Majee, P.; Sudhakar, S.; Mishra, S.; Kalia, J.; Pradeepkumar, P. I. and Srivatsan, S. G. Chem. Sci., 2024,15, 7982-7991. https://pubs.rsc.org/en/content/articlelanding/2024/sc/d4sc01755b
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