Journal Article

PUBDB-2025-05269

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png Live‐Cell RNA Imaging via Clickable Tri PPP ro Nucleotide Reporters

Knaack, J. I. H. ; List, E. ; Stalling, D.Extern*CSSB* ; Sterrenberg, V. T. ; Meier, C.Extern* ; Wagenknecht, H.-A. (Corresponding author) ; Bosse, J. B. (Corresponding author)Extern*CSSB*

2025
Wiley-VCH Weinheim

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Please use a persistent id in citations: doi:10.1002/anie.202516613  doi:10.3204/PUBDB-2025-05269

Abstract: Understanding RNA synthesis and dynamics in cells requires efficient labeling strategies that are not only compatible with cellular environments but can be performed in living cells. We developed a robust, bio-orthogonal approach for live-cell RNA labeling using TriPPPro (triphosphate prodrug) chemistry. This strategy enables the intracellular delivery of sterically demanding nucleoside triphosphates modified with inverse electron-demand Diels–Alder (IEDDA)-reactive groups, specifically trans-cyclooctene (2TCOa) and bicyclo[6.1.0]nonyne (BCN). Once hydrolyzed inside cells, these TriPPPro-modified uridines and cytidines are metabolically incorporated into nascent RNA by endogenous RNA polymerases. Subsequent IEDDA reaction with a dual-fluorogenic tetrazine-cyanine styryl dye conjugate allows wash-free, high-contrast imaging of RNA synthesis in cells. We demonstrate efficient RNA labeling, including nucleolar localization and specificity for newly transcribed RNA, validated by transcriptional inhibition and colocalization with ribosomal RNA. Comparative analyses confirm that TriPPPro delivery surpasses conventional transporter-based systems in both labeling efficiency and cellular compatibility. This platform offers a modular, non-genetic, and highly specific method for real-time RNA imaging, with broad applicability for RNA biology and antiviral research.

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Contributing Institute(s):

1. CSSB-MHH-JB (CSSB-MHH-JB)

Research Program(s):

1. 899 - ohne Topic (POF4-899) (POF4-899)
2. DFG project G:(GEPRIS)512741711 - SFB 1648: Neu auftretende Viren: Pathogenese, Struktur und Immunität (512741711) (512741711)

Experiment(s):

1. (Advanced Light and Fluorescence Microscopy Facility)

Appears in the scientific report 2025

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Database coverage:
; ; ; Chemical Reactions ; Clarivate Analytics Master Journal List ; Current Contents - Life Sciences ; Current Contents - Physical, Chemical and Earth Sciences ; DEAL Wiley ; Ebsco Academic Search ; Essential Science Indicators ; IF >= 15 ; Index Chemicus ; JCR ; Nationallizenz ; SCOPUS ; Science Citation Index Expanded ; Web of Science Core Collection

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