Targeting structured RNAs using cross-chiral recognition
RNA is now widely recognized to modulate a variety of cellular outcomes beyond serving as a template for protein expression. As with other biomolecules, the function of RNA is closely related to its three-dimensional structure. Both secondary and tertiary structural motifs serve as important recognition elements for RNA-RNA and RNA-protein interactions. In addition, structured RNA elements play critical roles in a variety of diseases, including viral infections, cancer, and neurological disorders. Despite having a well-recognized therapeutic and diagnostic potential, structured RNAs remain underutilized targets for disease intervention (outside of antibiotics targeting the ribosome). Therefore, the development of new strategies for targeting structured RNAs is of the utmost importance.
The goal of this research project is to develop L-RNA-based affinity reagents and catalysts for practical biomedical applications. Our approach utilizes “cross-chiral” recognition, which we define as the hybridization independent recognition that occurs between two nucleic acids of opposite stereochemistry. Using in vitro selection techniques, we have prepared cross-chiral aptamers comprised of L-RNA (the nuclease resistant enantiomer of natural D-RNA) that bind structured D-RNA targets based on their unique shape rather than primary sequence. Having demonstrated the therapeutic potential of cross-chiral aptamers, we are now focused on improving their properties using a variety of approaches, including the use of unnatural nucleotides. In addition, we are using both biochemical and structural approaches to investigating how nucleic acids of opposite stereochemistry interact. Finally, we are utilizing cross-chiral recognition to develop biosensors capable of detecting dynamic RNA modification in real-time.
Construction and analysis of “designer” chromatin containing specific DNA lesions.
The genome of living organisms is constantly damaged by exogenous and endogenous sources. If not properly repaired, damaged DNA can induce mutations, the accumulation of which leads to cancer and aging. Although dysregulation of DNA repair processes drives genome instability and ultimately disease progression, it also provides a therapeutic opportunity. A better understanding of DNA repair mechanisms will not only increase our knowledge of disease development, but also help us refine treatment.
The overarching goal of this research project is to integrate chemical, biochemical and biophysical tools for the purpose of providing a deeper understanding of the molecular mechanisms underlying the regulation of DNA repair within chromatin. Our approach relies on the in vitro construction of “designer” chromatin, which contain site-specific DNA and protein modifications. This approach allows precise control over the environment of a particular DNA lesion, enabling detailed analysis of the repair process in a variety of chromatin architectures. Emphasis is being placed on the process of chromatin remodeling resulting from both covalent histone modification, such as poly(ADP-ribosyl)ation, and ATP-dependent chromatin remodelers. Because very little is known about the mechanisms of chromatin remodeling in the context of DNA repair, this project will result in a much deeper understanding of how specific DNA lesions, such as those implicated in cancer and aging, are repaired in chromatin. In addition, DNA repair-specific factors identified by our studies may offer novel targets for disease diagnostics and/or therapeutics.