Abstract:
Eukaryotic cells have 2 to 3 discrete nucleoli required for ribosome synthesis. Nucleoli are phase separated nuclear sub-organelles. Here we examined the role of nuclear Lamins and nucleolar factors in modulating the compartmentalization and dynamics of histone 2B (H2B-ECFP) in the nucleolus. Live imaging and Fluorescence Recovery After Photobleaching (FRAP) of labelled H2B, showed that the depletion of Lamin B1, Fibrillarin (FBL) or Nucleostemin (GNL3), enhances H2B-ECFP mobility in the nucleolus. Furthermore, Nucleolin knockdown significantly decreases H2B-ECFP compartmentalization in the nucleolus, while H2B-ECFP residence and mobility in the nucleolus was prolonged upon Nucleolin overexpression. Co-expression of N-terminal and RNA binding domain (RBD) deletion mutants of Nucleolin or inhibiting 45S rRNA synthesis reduces the sequestration of H2B-ECFP in the nucleolus. Taken together, these studies reveal a crucial role of Nucleolin-rRNA complex in modulating the compartmentalization, stability and dynamics of H2B within the nucleolus.