Pre-rRNA spatial distribution and functional organization of the nucleolus.

Abstract

The multilayered nucleolus is the primary site of ribosome biogenesis^1,2, where successive maturation of small (SSU)^3,4 and large (LSU)⁵ ribosomal subunit precursors occurs. However, the spatiofunctional relationship between pre-rRNA processing and nucleolar substructures and how this adapts to changing cellular physiological demands have remained incompletely understood^6,7. Here our spatiotemporal analyses revealed a compartment-specific ribosomal subunit processing in human nucleoli, with SSU processomes maintained in fibrillar centre (FC)-dense fibrillar component (DFC)-periphery dense fibrillar component (PDFC) domains while LSU pre-rRNAs largely transited to PDFC-granular component regions. Slowly proliferating cells exhibited unexpected 5' external transcribed spacer (5' ETS)-centred SSU processing impairment, accompanied by structural remodelling of FC-DFC units and retarded SSU outflux. Direct 5' ETS processing perturbation at least partially recapitulated these FC-DFC unit alterations, supporting the functional interdependence between SSU processing and nucleolar architecture. Notably, anamniote bipartite nucleoli with merged FC-DFC compartments^8,9 exhibited distinct 5' ETS distribution and slower pre-rRNA flux compared with multilayered nucleoli in amniotes. Introducing a FC/DFC interface to bipartite nucleoli enhanced processing efficiency, indicating that the evolutionary emergence of nested FC-DFC units may have optimized pre-rRNA processing. Collectively, depicting the spatiotemporal distribution of pre-rRNAs reveals an essential role of 5' ETS-centred processing in maintaining nucleolar substructures and suggests a possible evolutionary advantage of the multilayered structure in amniotes.