In eukaryotic cells, the nucleolus is the central site for ribosome biogenesis. It is composed of distinct subcompartments: fibrillar center (FC), dense fibrillar component (DFC), periphery of DFC (PDFC), and granular component (GC), each executing specific steps in pre-rRNA transcription, processing, and ribosomal subunit assembly.
Nucleolar dysfunction is implicated in various diseases such as cancer, neurodegeneration, and developmental disorders. The traditional view of ribosome biogenesis process is that ribosomal RNA (rRNA) synthesis and processing occur in FC-DFC units, and ribosome assembly occurs in GC, but how these compartments functionally coordinate to support efficient ribosome production remains unknown.
In a study published in Nature on July 23, a team led by Prof. CHEN Lingling from the Center for Excellence in Molecular Cell Science (Shanghai Institute of Biochemistry and Cell Biology) of the Chinese Academy of Sciences revealed a spatiotemporal separation in the processing of distinct rRNA precursors (pre-rRNAs) within the nucleolus, offering a new perspective on how nucleolar architecture is functionally organized.
By integrating metabolic labeling, single-molecule RNA imaging, super-resolution microscopy, and quantitative proteomics, researchers systematically mapped the spatiotemporal distribution of pre-rRNA processing in the nucleolus.
They uncovered that small subunit (SSU) pre-rRNA was predominantly processed within the inner FC-PDFC regions and largely completed its processing within the first 30 minutes. While the large subunit (LSU) pre-rRNA was enriched in the outer PDFC-GC regions and matured gradually after 30 minutes. This spatial separation contradicts traditional models that placed both processing steps within the GC, and redefines the specialized functions within nucleolar subdomains.
Such a compartmentalized processing is functionally significant. In slow-proliferation or differentiated cells, nucleoli exhibited fewer but enlarged FC-DFC units, with accompanying less efficient SSU processing and accumulated SSU pre-rRNAs near the inner nucleolar regions, ultimately resulting in the reduced production of ribosomes.
To quantify the relationship between pre-rRNA processing and nucleolar organization, researchers introduced a new geometric parameter, named Relative FC/DFC interface, which quantitatively links individual nested FC/DFC structures with the SSU processing efficiency. Antisense oligonucleotides targeting the 5' external transcribed space (5' ETS) region of SSU pre-rRNA were used to inhibit pre-rRNA processing, recapitulating the structural defects observed in slow-proliferation cells. The findings revealed the essential role of 5' ETS-centered SSU processing in maintaining nucleolar substructures.
Notably, evolutionary comparisons of pre-rRNA processing kinetics further supported the model that the nucleolar structure and function are interdependent. Bipartite nucleoli in anamniotes such as zebrafish, which lacks a separated FC-DFC interface, exhibited distinct 5′ ETS distribution and much slower pre-rRNA diffusion compared to multilayered nucleoli in amniotes. The introduction of an artificial FC/DFC interface to bipartite nucleoli led to enhanced processing efficiency, suggesting that the emergence of multilayered nucleolar organization may have conferred evolutionary advantages in ribosome production.
This study establishes a direct link between molecular-level function and micron-level nucleolar architecture, revealing the dynamic coordination between pre-rRNA processing and nucleolar substructure, and offering potential strategies for nucleolus-related disease treatment via targeting key steps of ribosome biogenesis within nucleolar substructures.
