Chemical-guided SHAPE sequencing (cgSHAPE-seq) informs the binding site of RNA-degrading chimeras targeting SARS-CoV-2 5' untranslated region

  • Nat Commun. 2025 Jan 8;16(1):483. doi: 10.1038/s41467-024-55608-w.
Zhichao Tang  #  1  2 ,  Shalakha Hegde  #  1  2 ,  Siyuan Hao  3 ,  Manikandan Selvaraju  1 ,  Jianming Qiu  3 ,  Jingxin Wang  4  5
Affiliations
  • 1. Department of Medicinal Chemistry, University of Kansas, Lawrence, USA.
  • 2. Section of Genetic Medicine, Department of Medicine, Biological Sciences Division, University of Chicago, Chicago, USA.
  • 3. Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, USA.
  • 4. Department of Medicinal Chemistry, University of Kansas, Lawrence, USA. [email protected].
  • 5. Section of Genetic Medicine, Department of Medicine, Biological Sciences Division, University of Chicago, Chicago, USA. [email protected].
  • # Contributed equally.
PMID: 39779694 DOI: 10.1038/s41467-024-55608-w
Abstract

One of the hallmarks of RNA viruses is highly structured untranslated regions (UTRs) which are often essential for viral replication, transcription, or translation. In this report, we discovered a series of coumarin derivatives that bind to a four-way RNA helix called SL5 in the 5' UTR of the SARS-CoV-2 RNA genome. To locate the binding site, we developed a sequencing-based method namely cgSHAPE-seq, in which an acylating probe was directed to crosslink with the 2'-OH group of ribose at the binding site to create read-through mutations during reverse transcription. cgSHAPE-seq unambiguously determined a bulged G in SL5 as the primary binding site, which was validated through mutagenesis and in vitro binding experiments. The coumarin derivatives were further used as a warhead in designing RNA-degrading chimeras to reduce viral RNA expression levels. The optimized RNA-degrading chimera C64 inhibited live virus replication in lung epithelial carcinoma cells.

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