The Piccirilli group develops and uses chemical and biochemical tools to investigate the structure and function of noncoding RNAs (ncRNA) and their complexes with proteins (RNPs) and small molecules (smRNAs). In the context of this project, we will specifically focus on two general areas: 1) engineering fragment antigen binding (Fabs) as chaperones for RNA/ crystallography, cryoelectron microscopy (cryoEM) and as enabling tools for RNA biology, and 2) elaborating mechanisms of RNA function, especially catalytic RNAs (ribozymes). The terrestrial transcriptome contains vast numbers of structured RNAs within both coding and noncoding transcripts that act alone and in concert with proteins to carry out critically important roles in nearly every facet of cellular function. Understanding how these RNAs mediate biological function in health and disease requires knowledge of their three-dimensional architectures. We deploy a variety of structural biology, biophysical, biochemical, computational and chemical approaches, including kinetic isotope effect analysis and the use of nucleotides from an artificially expanded genetic system. Beyond the conceptual advances that emerge, our work delivers powerful Fab reagents for RNA/RNP structural and functional studies, nucleotide analogs to support novel approaches to define RNA catalytic mechanisms, and strategies to render large RNAs more tractable for study.
Engineering Antibody Fragments That Bind RNA and Facilitate Crystallization
The biomedical and biotechnology research communities have built a vast commercial and research infrastructure centered around antibody technology, as these versatile components of the immune system serve can serve as drugs, affinity reagents for cell biology, and as crystallization chaperones for structural biology. However, much of this powerful technology remains orthogonal to the investigation of RNA biology because traditional in vivo methods for generating antibodies are typically not amenable to RNA targets. To circumvent this limitation, we have pioneered the use of recombinant antibody technology for engineering antigen binding fragments (Fabs) against RNA. These efforts have engendered a new field with important applications in RNA structural biology, drug discovery, and delineating RNA processes in the cell.
RNA Catalysis
Beyond the well-recognized roles for RNA in nearly all aspects of protein synthesis, RNA engages in a diverse array of other regulatory functions via equally diverse mechanisms that operate to control genome dynamics, metabolism, developmental programming, and pathogenesis. Analysis of entire transcriptomes in the past decade has revealed the presence of vast numbers of these so-called non-coding RNAs, and this list and associated functions continues to grow rapidly. Natural ribozymes constitute a prominent class of non-coding RNAs that mediate diverse biological functions, and our work has helped to establish general paradigms for their catalytic mechanisms: (1) that RNA splicing machineries (group I and group II self-splicing introns and the spliceosome) use metal ions directly in catalysis, and (2) that small endonucleolytic ribozymes can use their own nucleobases to mediate general acid/base catalysis. A defining feature of our work is the combination of rigorous quantitative analysis with nucleic acid chemistry. Our mechanistic experience together with our ability to design and construct nucleoside analogues has allowed us to answer fundamental questions of RNA structure and function on a deeper level than would be possible using only the four natural nucleotides. In addition to revealing fundamental principles governing RNA catalysis, our work has engendered new approaches for investigating the chemical biology of RNA. These approaches are generally applicable to other non-catalytic RNAs (ncRNAs) and to protein enzymes that operate on nucleic acids.
Chemical Biology of RNA
The terrestrial transcriptome contains vast numbers of structured RNAs within both coding and noncoding transcripts that act alone and in concert with proteins to carry out critically important roles in nearly every facet of cellular function. Understanding how these RNAs mediate biological function in health and disease requires knowledge of their three-dimensional architectures. We deploy a variety of structural biology, biophysical, biochemical, computational and chemical approaches including RNA modification reagents, single-atom perturbations, and unnatural base pairs to deduce cellular roles.
University of Scranton
B.Sc.
1982
Fulbright Scholar, 1982-1983
1983
Harvard University,
Ph.D.
1989
Harvard Traveling Scholar
1989
Howard Hughes Postdoctoral Fellow
University of Colorado, Boulder
1993
University of Colorado at Boulder
Howard Hughes Postdoctoral Research Fellow
1993
The University of Chicago
Assistant Professor
2000
The University of Chicago
Professor
Present
Radakovic A, Lewicka A, Todisco M, Aitken HRM, Weiss Z, Kim S, Bannan A, Piccirilli JA, Szostak JW. A potential role for RNA aminoacylation prior to its role in peptide synthesis. Proc Natl Acad Sci U S A. 2024;121(35):e2410206121. Epub 20240823. doi: 10.1073/pnas.2410206121. PubMed PMID: 39178230; PMCID: PMC11363276.
Wilson TJ, McCarthy E, Ekesan S, Giese TJ, Li NS, Huang L, Piccirilli JA, York DM, Lilley DMJ. The Role of General Acid Catalysis in the Mechanism of an Alkyl Transferase Ribozyme. ACS Catal. 2024;14(20):15294-305. Epub 20241002. doi: 10.1021/acscatal.4c04571. PubMed PMID: 39444533; PMCID: PMC11494507.
Krochmal D, Roman C, Lewicka A, Shao Y, Piccirilli JA. Structural basis for promiscuity in ligand recognition by yjdF riboswitch. Cell Discov. 2024;10(1):37. Epub 20240402. doi: 10.1038/s41421-024-00663-2. PubMed PMID: 38565535; PMCID: PMC10987639.
Lewicka A, Roman C, Jones S, Disare M, Rice PA, Piccirilli JA. Crystal structure of a cap-independent translation enhancer RNA. Nucleic Acids Res. 2023;51(16):8891-907. doi: 10.1093/nar/gkad649. PubMed PMID: 37548413; PMCID: PMC10484670.
Yoon S, Ollie E, York DM, Piccirilli JA, Harris ME. Rapid Kinetics of Pistol Ribozyme: Insights into Limits to RNA Catalysis. Biochemistry. 2023;62(13):2079-92. Epub 20230609. doi: 10.1021/acs.biochem.3c00160. PubMed PMID: 37294744; PMCID: PMC10330772.
Weissman B, Ekesan S, Lin HC, Gardezi S, Li NS, Giese TJ, McCarthy E, Harris ME, York DM, Piccirilli JA. Dissociative Transition State in Hepatitis Delta Virus Ribozyme Catalysis. J Am Chem Soc. 2023;145(5):2830-9. Epub 20230127. doi: 10.1021/jacs.2c10079. PubMed PMID: 36706353; PMCID: PMC10112047.
Roman C, Lewicka A, Koirala D, Li NS, Piccirilli JA. The SARS-CoV-2 Programmed -1 Ribosomal Frameshifting Element Crystal Structure Solved to 2.09 A Using Chaperone-Assisted RNA Crystallography. ACS Chem Biol. 2021;16(8):1469-81. Epub 20210730. doi: 10.1021/acschembio.1c00324. PubMed PMID: 34328734; PMCID: PMC8353986.
Ganguly A, Weissman BP, Giese TJ, Li NS, Hoshika S, Rao S, Benner SA, Piccirilli JA, York DM. Confluence of theory and experiment reveals the catalytic mechanism of the Varkud satellite ribozyme. Nat Chem. 2020;12(2):193-201. Epub 20200120. doi: 10.1038/s41557-019-0391-x. PubMed PMID: 31959957; PMCID: PMC7389185.
Investigator,
Howard Hughes Medical Institute
1994 - 2009
The Llewellyn John and Harriet Manchester Quantrell Award for Excellence in Undergraduate Teaching
1998
Chair
National Institutes of Health Study Section: Macromolecular Structure and Function A
2017 - 2018
Editorial Board
RNA Journal
2018 - 2022