Structural investigation of an RNA device that regulates <i>PD-1</i> expression in mammalian cells

Synthetic RNA devices are engineered to control gene expression and offer great potential in both biotechnology and clinical applications. Here, we present multidisciplinary structural and biochemical data for a tetracycline (Tc)-responsive RNA device (D43) in both ligand-free and bound states, providing a structure-dynamical basis for signal transmission. Activation of self-cleavage is achieved via ligand-induced conformational and dynamical changes that stabilize the elongated bridging helix harboring the communication module, which drives proper coordination of the catalytic residues. We then show the utility of CRISPR-integrated D43 in EL4 lymphocytes to regulate programmed cell death protein 1 (PD-1), a key receptor of immune checkpoints. Treatment of these cells with Tc showed a dose-dependent reduction in PD-1 by immunostaining and a decrease in messenger RNA levels by quantitative PCR as compared with wild type. PD-1 expression was recoverable upon removal of Tc. These results provide mechanistic insight into RNA devices with potential for cancer immunotherapy or other applications.

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Publication Details

Journal
Nucleic Acids Research
Published
2025-02-27
DOI
https://doi.org/10.1093/nar/gkaf156
Primary Topic
RNA Interference and Gene Delivery
Type
article
Field-Weighted Citation Impact
0.00

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article

Structural investigation of an RNA device that regulates PD-1 expression in mammalian cells

Szu-Yun Chen, Yun‐Xing Wang, Yun-Tzai Lee, Raj Chari et al.
Nucleic Acids Research
RNA Interference and Gene Delivery
article

Structural investigation of an RNA device that regulates PD-1 expression in mammalian cells

Szu-Yun Chen, Yun‐Xing Wang, Yun-Tzai Lee, Raj Chari, Susan M. Lea, Vibha Dwivedi, Justin C. Deme, Maximilia Frazão de Souza Degenhardt, Jason R. Stagno, Howard A. Young, Lixin Fan, Hyun Kyung Lee, Ping Yu
article en

Abstract

Synthetic RNA devices are engineered to control gene expression and offer great potential in both biotechnology and clinical applications. Here, we present multidisciplinary structural and biochemical data for a tetracycline (Tc)-responsive RNA device (D43) in both ligand-free and bound states, providing a structure-dynamical basis for signal transmission. Activation of self-cleavage is achieved via ligand-induced conformational and dynamical changes that stabilize the elongated bridging helix harboring the communication module, which drives proper coordination of the catalytic residues. We then show the utility of CRISPR-integrated D43 in EL4 lymphocytes to regulate programmed cell death protein 1 (PD-1), a key receptor of immune checkpoints. Treatment of these cells with Tc showed a dose-dependent reduction in PD-1 by immunostaining and a decrease in messenger RNA levels by quantitative PCR as compared with wild type. PD-1 expression was recoverable upon removal of Tc. These results provide mechanistic insight into RNA devices with potential for cancer immunotherapy or other applications.

Nucleic Acids ResearchVol. 53(5)
Frederick National Laboratory for Cancer Research (US), National Cancer Institute (US), Center for Cancer Research (US)
U.S. Department of Health and Human Services, National Institutes of Health, National Cancer Institute, National Institute of General Medical Sciences, Basic Energy Sciences, Biological and Environmental Research
Good health and well-being
Openalex Percentile: Top 96%
RNA Interference and Gene Delivery
DOIOpen Access PDFOpenAlex
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