Sunday, May 18, 2025
5/18/2025
Molecular mechanisms that regulate ADAR target recognition and RNA editing - PROJECT SUMMARY/ABSTRACT RNA-protein interactions underlie proper control of gene expression across all domains of life. Defining structural elements and sequences recognized by RNA binding proteins is critical to understand both the biological function and pathological consequences of dysregulated interactions. Viral, prokaryotic, and eukaryotic RNA binding proteins that recognize double-stranded RNA (dsRNA) impact nearly all aspects of RNA metabolism, yet a molecular understanding of how these proteins recognize specific cellular targets is lacking. This Maximizing Investigators Research Award application is proposed to support research in the Hundley lab focused on connecting the molecular mechanisms of dsRNA recognition by the ADAR family of RNA binding proteins to functional consequences on gene expression that impact development and disease. ADARs bind dsRNA and catalyze the deamination of adenosine to inosine in mRNA, both of which are essential functions of human ADARs and dysregulated in over 35 human diseases. The ability of ADARs to change the genome-encoded information present in RNA provides an important means to diversify the transcripts expressed in an organism’s tissues over time and is being harnessed for personalized medicine approaches to correct mutations at the RNA level and improve human health. However, factors that control ADAR function in vivo are poorly understood. The proposed work focuses on fundamental questions of how ADAR binding to target RNAs is influenced by transcription and cellular metabolism, and how these events influence RNA fate to impact neuronal function. To answer these broad questions, we are taking an integrated approach using human cell lines and Caenorhabditis elegans, the latter of which provides an important platform for mechanistic discovery as RNA binding and deamination are controlled by different proteins. Biochemical and transcriptome-wide discovery of binding sites of these two proteins coupled with in vivo model testing and new efforts in structural biology will afford a unique opportunity to reveal how binding and deamination are coordinated. Furthermore, the use of a genetically tractable multicellular organism amenable to behavioral assays and neural-specific transcriptomics allows us to connect consequences of ADAR binding and editing to neuronal function and organismal physiology. Unbiased and genome-wide approaches will facilitate identification of novel cellular factors that influence the molecular signature of ADAR binding. Work in cell lines will be centered on ADAR3, a human deaminase-deficient ADAR protein that regulates editing in glioblastoma (brain cancer). New research will focus on dissecting the mechanistic connections between altered editing, RNA binding and immune activation by ADAR3. Overall, our work will define mechanism that regulate ADAR activity and provide possible targets for therapeutic development, in addition to fundamentally advancing the fields of RNA editing and dsRNA-mediated cellular pathways.
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