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Lead-seq: transcriptome-wide structure probing in vivo using lead(II) ions.

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Researchers developed Lead-seq, a new method to map RNA structures in living cells. This technique revealed temperature-dependent RNA structures in the pathogen Yersinia pseudotuberculosis, offering insights into gene regulation.

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Area of Science:

  • Molecular Biology
  • Genomics
  • Biophysics

Background:

  • Understanding RNA structure in vivo is crucial for deciphering gene function.
  • Existing methods like SHAPE, kethoxal, and DMS have advanced RNA structuromics.
  • A need exists for methods providing base-independent structural information.

Purpose of the Study:

  • To establish and validate lead probing as a global RNA structuromics approach.
  • To investigate the transcriptome-wide RNA landscape of Yersinia pseudotuberculosis.
  • To identify temperature-responsive RNA structures and their regulatory roles.

Main Methods:

  • Development of Lead-seq: combining lead(II) acetate-mediated RNA cleavage with high-throughput sequencing.
  • Application of Lead-seq to Yersinia pseudotuberculosis under varying temperature conditions.
  • Validation of Lead-seq by recapitulating known RNA secondary structures.

Main Results:

  • Lead-seq provides RNA structural information independent of base identity.
  • The method successfully mapped secondary structures of various RNAs, including tRNAs, rRNAs, and mRNAs.
  • The first temperature-responsive in vivo RNA structurome of a bacterial pathogen was unveiled.
  • Temperature-regulated translation of candidate genes identified through Lead-seq was confirmed.

Conclusions:

  • Lead-seq is a valuable complementary approach for interrogating intracellular RNA structures globally.
  • The study provides novel insights into the dynamic RNA structurome of Yersinia pseudotuberculosis.
  • Lead-seq facilitates the discovery of functionally relevant, environmentally responsive RNA structures.