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Intrinsically disordered regions (IDRs) in Pseudomonas aeruginosa RNA helicases RhlE1 and RhlE2 dictate their distinct functions by modulating RNA binding, phase separation, and interactions with other cellular components like RNase E.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Microbiology

Background:

  • RNA helicases are crucial for RNA metabolism, often possessing intrinsically disordered regions (IDRs) that facilitate complex interactions and phase separation.
  • Pseudomonas aeruginosa utilizes two non-redundant RNA helicases, RhlE1 and RhlE2, which share a catalytic core but differ in their C-terminal IDRs.

Purpose of the Study:

  • To investigate how variations in intrinsically disordered regions (IDRs) between RhlE1 and RhlE2 RNA helicases confer functional specificity.
  • To elucidate the roles of IDRs in RNA binding, phase separation, enzymatic activity, and interactions with other cellular factors.

Main Methods:

  • Construction and analysis of chimeric RNA helicases with swapped IDRs.
  • Biochemical assays to assess RNA unwinding activity.
  • Analysis of protein-protein interactions, specifically with RNase E.
  • Assessment of bacterial growth under varying temperature conditions.
  • Localization studies of RNA helicases within the cytoplasm.

Main Results:

  • Both RhlE1 and RhlE2 IDRs promote RNA binding and cytoplasmic phase separation.
  • The RhlE2 IDR enhances RNA unwinding activity and phase separation more effectively than the RhlE1 IDR.
  • The RhlE2 IDR mediates interaction with the RNase E endonuclease.
  • Chimeric helicases exhibit distinct biochemical and functional properties compared to native forms.
  • The RECRhlE1-IDRRhlE2 chimera improved cold growth and interacted with RNase E.
  • The RECRhlE2-IDRRhlE1 chimera impaired bacterial growth at low temperatures, linked to aberrant RNA droplets.

Conclusions:

  • Intrinsically disordered regions (IDRs) are key determinants of functional specificity in RNA helicases.
  • IDRs modulate both the catalytic core activity and the subcellular localization of RNA helicases through phase separation.
  • Understanding IDR diversity is essential for deciphering the functional differentiation of RNA helicases in bacteria like Pseudomonas aeruginosa.