RNA BioChemistry

Faculty currently associated with this research theme.

Dissecting RNA structure, dynamics and function using novel methods of simulatio +

Impact: The recent completion of the ENCODE (Encyclopedia of DNA elements) project has brought to light the importance of RNA in cellular processes; a significant portion of the human genome is transcribed as non-coding RNA and these molecules play a critical role in genetic regulation, catalysis and are linked to many diseases. Moreover, there is growing understanding of the role pH plays in modulating the structure and function of molecules, including RNA, within the cell. The development of RNA targeted drugs, the understanding of catalytic processing and conformational switching by RNA all require an integrated understanding of the structure and dynamics of RNA within the pH environments of the cell.

Objective: To develop and implement a theoretical and computational framework to explore and understand RNA structure and dynamics and the role pH plays in modulating and controlling chemical and physical processes of these molecules in the cell.

Approach: Our group has recently developed novel methods to integrate the influence of pH into molecular simulations, thereby enabling the thorough study of pH-mediated processes in RNA molecules. Additionally, we have developed, implemented and are applying methods of structure prediction and free energy landscape exploration for RNA folding and function. Our integrated approach brings together rigorous statistical mechanical theory and molecular simulations approaches that we are utilizing in collaborative studies to understand aspects of protein synthesis by the ribosome, ribosome assembly, conformational switching in translation and transcription and the role of pH in ribozyme function.

Charles Brooks

Dissecting RNA Mechanisms inside Live Cells at the Single Molecule Level +

Impact: Since only the turn of the millennium, small non-coding RNAs have rapidly emerged as a central class of regulators of eukaryotic gene expression, affecting all aspects of multi-cellular life. The diverse class of small RNAs includes microRNAs (miRNAs) and small interfering RNAs (siRNAs), both of which assemble with protein components into RNA induced silencing complexes (RISC) that down-regulate RNA transcripts. To date, almost 1,500 unique mammalian miRNAs have been identified that, collectively, represent over 2% of the human genome and are predicted to regulate over 60% of all protein coding genes in a complex network of interactions in which they bind to the 3’ untranslated regions (UTRs) of messenger (m)RNAs to repress protein expression via RNA silencing. By contrast, siRNAs are directly derived through cleavage from long stretches of double-stranded RNAs associated with viruses and transposable elements, and lead to cleavage of the target virus or transposon RNA in a process referred to as RNA interference. The prospect of using small RNAs to control gene expression has spawned a multi-billion dollar industry, aiming to develop therapies against a host of human diseases.

Objective: To detect single small RNA molecules such as miRNAs and siRNA inside living cells as they go about their biological functions in their “natural habitat”.

Approach: We recently developed iSHiRLoC, or intracellular Single molecule, High-Resolution Localization and Counting, as an innovative probe concept optimized for detecting single small RNA molecules inside living cells. For iSHiRLoC we microinject fluorophore labeled RNAs into cultured human cells at low enough numbers not to overwhelm the cellular machinery. We then either track single RNA-containing particles diffusing in the cell at super-resolution or count the number of RNA molecules they contain by stepwise photobleaching of the attached fluorophores.

Nils Walter

Additional RNA Biochemistry projects

Additional projects are focused on the structural and mechanistic bases of HIV-1 RNA dependent replication functions at the cellular, viral and atomic levels (Hashim Al-Hashimi) and on developing biophysical methods, including single molecule fluorescence and NMR spectroscopy, to investigate hallmark features of large RNA molecules such as RNase P, responsible for processing of all tRNAs (Carol Fierke).