Non-coding RNAs (ncRNAs) represent a vast and largely untapped dimension of biology, accounting for more than 90% of the transcriptional output of the human genome. This challenges earlier notions that much of our DNA is mere “junk.” With over 80,000 unique ncRNAs identified in human cells, these molecules play intricate roles in genome maintenance, processing, and regulation. Nature, akin to modern nanotechnology, leverages RNA to create nanoscale machines with remarkable architectural and functional complexity. We are using advanced fluorescence microscopy techniques, such as single-molecule fluorescence resonance energy transfer (smFRET) and super-resolution imaging, for probing these RNA structures in real-time with nanometer precision.
Among these captivating ncRNA elements are riboswitches—highly structured RNA elements embedded in the 5’ untranslated regions of bacterial mRNAs. These motifs regulate gene expression by folding co-transcriptionally to bind small molecules or ions, acting as precise molecular switches. For instance, manganese-sensing riboswitches utilize a central, adaptable RNA helix as a molecular fulcrum, integrating diverse signals to achieve finely tuned bacterial gene regulation. The Mn2+ riboswitch exemplifies a key feature of riboswitch architecture: while its upstream aptamer region is highly conserved to ensure robust ligand binding, the downstream expression platform often exhibits lower conservation due to its species-specific interactions with protein effectors. This nuanced functional adaptability highlights the limitations of relying solely on sequence conservation to predict the functional importance of nucleotides within riboswitches.
Our research employs single-molecule imaging and biochemical techniques to reveal how RNA folding dynamically couples with gene expression regulation. Riboswitches, as we uncover, epitomize the intricate interplay between RNA structure and function, enabling exquisite regulatory precision. These findings illuminate the untapped potential of ncRNAs as vital players in life’s processes, expanding our understanding of biological regulation and paving the way for innovative therapeutic strategies.