New labeling technique reveals hidden structures of cancer-linked RNA molecules
Researchers have developed a chemical tagging method that makes RNA signals 27 times brighter under magnetic imaging, solving a decades-old obstacle in studying disease-causing genetic regulators. The breakthrough could accelerate drug discovery for cancer and other RNA-related diseases by enabling scientists to observe molecular behavior directly in cells for the first time.
Originaltitel: Targeted NMR signal enhancement of RNA by site-directed bis-nitroxide labeling
MicroRNAs regulate gene expression through sequence-specific interactions with target messenger RNAs (mRNAs), and their misregulation is a hallmark of cancer. MicroRNA-34a (miR-34a), a key modulator of the tumor suppressor p53, binds the mRNA encoding sirtuin 1 (mSirt1) and adopts multiple conformational states that influence repression efficiency. While such dynamics have been characterized in vitro, extending these studies to cellular environments is hampered by weak signals and substantial background inherent to nucleic acid NMR. To overcome this limitation, we developed a site-directed spin labeling strategy for RNA that enables targeted dynamic nuclear polarization (DNP) signal enhancement. Using the bisnitroxide polarizing agent AsymPol-NCS-SDSL, we conjugated spin labels to specific positions of mSirt1 RNA and annealed them to 13 C, 15 N-cytidine-labeled miR-34a. At 9.4 T, we observed up to 27-fold signal enhancements. The selectivity of polarization transfer within the RNA duplex relative to the surrounding environment could be tuned by matrix deuteration, while doping with paramagnetic metal ions accelerated polarization build-up times, with Cu II proving more efficient than Gd III . This work establishes bisnitroxide-based SDSL as a powerful approach for targeted DNP of nucleic acids, enabling high-sensitivity studies of nucleic acids at concentrations ≤40 µ m and paves the way for structural investigations of microRNA–mRNA interactions in cells.