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Chemotranscriptomic profiling with a thiamine monophosphate photoaffinity probe.

Stefan Crielaard1, Casper F M Peters1, Alexandar Slivkov1

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Researchers used a novel chemotranscriptomic method to discover new RNA-small molecule interactions in Escherichia coli. They identified Thiamine Monophosphate (TMP) interacting with a riboswitch, suggesting a potential biological role for TMP.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genomics

Background:

  • Riboswitches are RNA molecules that regulate gene expression by binding small molecules.
  • Understanding RNA-small molecule interactions is crucial for deciphering cellular regulatory networks.

Purpose of the Study:

  • To profile the Escherichia coli transcriptome for interactions with Thiamine Monophosphate (TMP).
  • To identify novel RNA-small molecule interactions using chemotranscriptomic profiling.

Main Methods:

  • Design and synthesis of a TMP-based photoaffinity probe.
  • Chemotranscriptomic profiling using next-generation RNA sequencing.
  • Validation of interactions using RT-qPCR, competition assays, and SHAPE experiments.

Main Results:

  • Identified potential interactions between the TMP probe and bacterial transcripts.
  • Validated a significant interaction between the TMP probe and the Flavin Mononucleotide (FMN) riboswitch.
  • Localized photocrosslinked nucleotides to the riboswitch aptamer and confirmed moderate binding of TMP.

Conclusions:

  • Chemotranscriptomic profiling is a viable method for discovering new RNA-small molecule interactions.
  • Thiamine Monophosphate (TMP) selectively binds to the FMN riboswitch, potentially influencing gene expression.
  • TMP may have a biological role distinct from its natural ligand, FMN.