Dynamics and compartmentation of the nucleolar processing machinery

Area of Science:

• Cell Biology
• Molecular Biology
• Biochemistry

Background:

• Active nucleoli compartmentalize ribosomal RNA (rRNA) biogenesis machinery.
• Late rRNA processing proteins reside in the granular component (GC).

Purpose of the Study:

• Investigate the behavior of late rRNA processing proteins when 28S rRNA production is impaired and restored.
• Determine factors influencing nucleolar protein compartmentation.

Main Methods:

• Utilized 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) to impair rDNA transcription and rRNA processing.
• Observed nucleolar reformation dynamics.
• Analyzed protein traffic using fluorescence recovery after photobleaching (FRAP).

Main Results:

• DRB treatment dispersed rDNA clusters and disconnected late rRNA processing proteins from transcription sites.
• Processing proteins formed independent masses lacking detectable 28S rRNA, showing compartmentation persists without active processing.
• Reversal of DRB treatment restored rRNA processing and nucleolar reformation in two temperature-sensitive, energy-dependent steps.
• FRAP analysis revealed similar protein dynamics in disconnected masses and active GC, suggesting protein interactions are key.

Conclusions:

• Nucleolar compartmentation of late rRNA processing machinery is independent of active rRNA processing.
• Nucleolar reformation involves dynamic, energy-dependent recruitment of proteins to transcription sites and rDNA compaction.
• Protein dynamics and interactions, rather than solely processing activity, govern the compartmentation of nucleolar machineries.