Structural insights into coordinating 5S RNP rotation with ITS2 pre-RNA processing during ribosome formation

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• The rixosome is a key complex involved in ribosome biogenesis and gene silencing in eukaryotes.
• Previous studies have defined its roles in yeast and human systems.

Purpose of the Study:

• To isolate and structurally characterize the conserved rixosome from the thermophilic fungus Chaetomium thermophilum.
• To elucidate the molecular mechanisms by which the rixosome coordinates pre-ribosomal RNA processing and ribosome maturation.

Main Methods:

• Cryo-electron microscopy (Cryo-EM) or X-ray crystallography to determine the structure of the rixosome.
• Biochemical assays to study the enzymatic activities of the rixosome sub-modules (Las1 endonuclease, Grc3 polynucleotide-kinase).
• In vitro reconstitution assays to demonstrate the functional coordination of the rixosome modules in pre-rRNA processing.

Main Results:

• The rixosome from C. thermophilum is composed of two distinct sub-modules: the Rix1-Ipi3-Ipi1 complex and the Las1-Grc3 complex, linked flexibly.
• The Rix1 complex positions the 5S rRNA for proper folding and maturation on pre-60S particles.
• The Las1-Grc3 module cleaves and 5' phosphorylates the internal transcribed spacer 2 (ITS2) region of pre-rRNA, facilitating its removal by the Mtr4 helicase-exosome pathway.

Conclusions:

• The determined structure reveals the intricate coordination between different rixosome modules in facilitating crucial steps of ribosome biogenesis.
• The findings provide mechanistic insights into how the rixosome orchestrates rRNA processing events, including ITS2 removal.
• The conserved nature of the rixosome suggests that similar structural and functional strategies may be employed in gene silencing pathways.