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The nucleolus is a membrane-less nuclear structure that disassembles when cells undergo mitosis. During mitosis, nucleolar factors are thus released from the nucleolus and dynamically change their subcellular localization; however, their functions remain largely uncharacterised. Here, we found that a nucleolar factor called nucleolar protein 11 (NOL11) forms a protein complex with two tryptophan-aspartic acid (WD) repeat proteins named WD-repeat protein 43 (WDR43) and Cirhin in mitotic cells. This complex, referred to here as the NWC (NOL11-WDR43-Cirhin) complex, exists in nucleoli during interphase and translocates to the periphery of mitotic chromosomes, i.e., perichromosomal regions. During mitotic progression, both the congression of chromosomes to the metaphase plate and sister chromatid cohesion are impaired in the absence of the NWC complex, as it is required for the centromeric enrichment of Aurora B and the associating phosphorylation of histone H3 at threonine 3. These results reveal the characteristics of a novel protein complex consisting of nucleolar proteins, which is required for regulating kinetochores and centromeres to ensure faithful chromosome segregation., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The perichromosomal layer (PCL) is a structure that surrounds mitotic chromosomes, found in both animal and plant cells. It comprises various proteins and RNAs, mainly derived from the nucleolus. Several functions for the PCL have been suggested; however, the mechanism of PCL organization during mitosis remains unclear. The localization of several nucleolar proteins to the PCL is reportedly dependent on pre-ribosomal RNAs and the marker of proliferation, Ki67, which is a major PCL-localized protein. Here we demonstrate that, although the removal of pre-ribosomal RNAs from the PCL causes PCL delocalization of several nucleolar proteins, it does not affect the localization of Ki67. Conversely, Ki67 depletion results in the dissociation of both pre-ribosomal RNAs and nucleolar proteins from the PCL, which indicates that Ki67 is required for the PCL accumulation of pre-ribosomal RNAs, to which several nucleolar proteins are associated. Given these findings, we propose a model for PCL organization that comprises three essential layers: the scaffolding protein Ki67, pre-ribosomal RNAs for linkage, and outer nucleolar proteins., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The nucleolus, whose primary function is ribosome biogenesis, plays an essential role in p53 activation. Ribosome biogenesis is inhibited in response to cellular stress and several nucleolar proteins translocate from the nucleolus to the nucleoplasm, where they activate p53. In this study, we analysed precisely how impaired ribosome biogenesis regulates the activation of p53 by depleting nucleolar factors involved in rRNA transcription or rRNA processing. Nucleolar RNA content decreased when rRNA transcription was inhibited. In parallel with the reduced levels of nucleolar RNA content, the nucleolar protein Myb-binding protein 1 A (MYBBP1A) translocated to the nucleoplasm and increased p53 acetylation. The acetylated p53 enhanced p21 and BAX expression and induced apoptosis. In contrast, when rRNA processing was inhibited, MYBBP1A remained in the nucleolus and nonacetylated p53 accumulated, causing cell cycle arrest at the G1 phase by inducing p21 but not BAX. We propose that the nucleolus functions as a stress sensor to modulate p53 protein levels and its acetylation status, determining cell fate between cell cycle arrest and apoptosis by regulating MYBBP1A translocation.

Various cellular stresses activate autophagy, which is involved in lysosomal degradation of cytoplasmic materials for maintaining nutrient homeostasis and eliminating harmful components. Here, we show that RNA polymerase I (Pol I) transcription inhibition induces nucleolar disruption and autophagy. Treatment with autophagy inhibitors or siRNA specific for autophagy-related (ATG) proteins inhibited autophagy but not nucleolar disruption induced by Pol I transcription inhibition, which suggested that nucleolar disruption was upstream of autophagy. Furthermore, treatment with siRNA specific for nucleolar protein nucleophosmin (NPM) inhibited this type of autophagy. This showed that NPM was involved in autophagy when the nucleolus was disrupted by Pol I inhibition. In contrast, NPM was not required for canonical autophagy induced by nutrient starvation, as it was not accompanied by nucleolar disruption. Thus, our results revealed that, in addition to canonical autophagy, there may be NPM-dependent autophagy associated with nucleolar disruption.

Tetramerization of p53 is crucial to exert its biological activity, and nucleolar disruption is sufficient to activate p53. We previously demonstrated that nucleolar stress induces translocation of the nucleolar protein MYBBP1A from the nucleolus to the nucleoplasm and enhances p53 activity. However, whether and how MYBBP1A regulates p53 tetramerization in response to nucleolar stress remain unclear. In this study, we demonstrated that MYBBP1A enhances p53 tetramerization, followed by acetylation under nucleolar stress. We found that MYBBP1A has two regions that directly bind to lysine residues of the p53 C-terminal regulatory domain. MYBBP1A formed a self-assembled complex that provided a molecular platform for p53 tetramerization and enhanced p300-mediated acetylation of the p53 tetramer. Moreover, our results show that MYBBP1A functions to enhance p53 tetramerization that is necessary for p53 activation, followed by cell death with actinomycin D treatment. Thus, we suggest that MYBBP1A plays a pivotal role in the cellular stress response.

Nucleolar dynamics are important for cellular stress response. We previously demonstrated that nucleolar stress induces nucleolar protein Myb-binding protein 1A (MYBBP1A) translocation from the nucleolus to the nucleoplasm and enhances p53 activity. However, the underlying molecular mechanism is understood to a lesser extent. Here we demonstrate that MYBBP1A interacts with lysine residues in the C-terminal regulatory domain region of p53. MYBBP1A specifically interacts with nonacetylated p53 and induces p53 acetylation. We propose that MYBBP1A dissociates from acetylated p53 because MYBBP1A did not interact with acetylated p53 and because MYBBP1A was not recruited to the p53 target promoter. Therefore, once p53 is acetylated, MYBBP1A dissociates from p53 and interacts with nonacetylated p53, which enables another cycle of p53 activation. Based on our observations, this MYBBP1A-p53 binding property can account for efficient p53-activation by MYBBP1A under nucleolar stress. Our results support the idea that MYBBP1A plays catalytic roles in p53 acetylation and activation., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background: Tumor suppressor p53 is mutated in a wide variety of human cancers and plays a critical role in anoikis, which is essential for preventing tumorigenesis. Recently, we found that a nucleolar protein, Myb-binding protein 1a (MYBBP1A), was involved in p53 activation. However, the function of MYBBP1A in cancer prevention has not been elucidated., Methods: Relationships between MYBBP1A expression levels and breast cancer progression were examined using patient microarray databases and tissue microarrays. Colony formation, xenograft, and anoikis assays were conducted using cells in which MYBBP1A was either knocked down or overexpressed. p53 activation and interactions between p53 and MYBBP1A were assessed by immunoprecipitation and western blot., Results: MYBBP1A expression was negatively correlated with breast cancer tumorigenesis. In vivo and in vitro experiments using the breast cancer cell lines MCF-7 and ZR-75-1, which expresses wild type p53, showed that tumorigenesis, colony formation, and anoikis resistance were significantly enhanced by MYBBP1A knockdown. We also found that MYBBP1A binds to p53 and enhances p53 target gene transcription under anoikis conditions., Conclusions: These results suggest that MYBBP1A is required for p53 activation during anoikis; therefore, it is involved in suppressing colony formation and the tumorigenesis of breast cancer cells. Collectively, our results suggest that MYBBP1A plays a role in tumor prevention in the context of p53 activation.