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Bacterial Rps3 counters oxidative and UV stress by recognizing and processing AP-sites on mRNA via a novel mechanism

Mohammad Afsar1, Ankita Shukla1, Faiz Ali1,2

  • 1Biochemistry and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow-226031, India.

Nucleic Acids Research
|November 26, 2024
PubMed

Insights

Prokaryotic ribosomes, specifically ribosomal protein Rps3, can detect and cleave damaged mRNA at AP-sites. This discovery reveals a novel mechanism for bacterial translation quality control, distinct from eukaryotic pathways.

Area of Science:

  • Molecular Biology
  • Structural Biology
  • Genetics

Background:

  • mRNA secondary structures and lesions impede translation, causing ribosome stalling.
  • Prokaryotic ribosomal proteins Rps3, Rps4, and Rps5 in the mRNA entry tunnel act as an mRNA helicase center.
  • The mechanism by which ribosomes detect mRNA lesions remains unclear.

Purpose of the Study:

  • To investigate the apurinic/apyrimidinic (AP) endoribonuclease activity of bacterial ribosomes on AP-site containing mRNA.
  • To elucidate the role of ribosomal protein Rps3 in recognizing and processing mRNA lesions.

Main Methods:

  • Cryo-electron microscopy (Cryo-EM) for structural analysis.
  • Biochemical assays to assess endoribonuclease activity.
  • Knockdown experiments in Mycobacterium tuberculosis to confirm in vivo function.

Main Results:

  • Biochemical assays identified the Rps3 130RR131 motif as crucial for AP-site recognition and cleavage.
  • Cryo-EM revealed cleaved mRNA within the 30S ribosome entry tunnel.
  • Knockdown studies demonstrated Rps3's protective role against oxidative and UV stress in M. tuberculosis.

Conclusions:

  • Prokaryotic ribosomal protein Rps3 possesses novel AP-endoribonuclease activity, distinct from eukaryotic mechanisms.
  • Rps3 plays a key role in bacterial mRNA quality control by processing AP-sites.
  • This mechanism contributes to cellular protection against DNA-damaging agents.

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