Nucleophosmin integrates within the nucleolus via multi-modal interactions with proteins displaying R-rich linear motifs and rRNA

Mitrea, Diana M.; Cika, Jaclyn A; Guy, Clifford S.; Ban, David; Banerjee, Priya R.; Stanley, Christopher B.; Nourse, Amanda; Deniz, Ashok A.; Kriwacki, Richard W.

doi:10.7554/elife.13571

Cited by 446 publications

(529 citation statements)

References 70 publications

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“…We conclude that the integrity of mature heterochromatin domains relies on weak hydrophobic interactions, and that dimerization and interactions with non-histone binding partners contribute to HP1 immobilization. This is consistent with evidence that networks of multivalent interactions promote demixing in vitro and in vivo 7,19 . We propose that mature heterochromatic domains consist of both immobile (static) and mobile (liquid) HP1a compartments, similar to recent findings for nucleoli 20 .…”

supporting

confidence: 91%

Phase separation drives heterochromatin domain formation

Strom

Emelyanov

Mir

et al. 2017

Nature

1,639

1,431

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Constitutive heterochromatin is an important component of eukaryotic genomes that has essential roles in nuclear architecture, DNA repair and genome stability1, and silencing of transposon and gene expression2. Heterochromatin is highly enriched for repetitive sequences, and is defined epigenetically by methylation of histone H3 at lysine 9 and recruitment of its binding partner heterochromatin protein 1 (HP1). A prevalent view of heterochromatic silencing is that these and associated factors lead to chromatin compaction, resulting in steric exclusion of regulatory proteins such as RNA polymerase from the underlying DNA3. However, compaction alone does not account for the formation of distinct, multi-chromosomal, membrane-less heterochromatin domains within the nucleus, fast diffusion of proteins inside the domain, and other dynamic features of heterochromatin. Here we present data that support an alternative hypothesis: that the formation of heterochromatin domains is mediated by phase separation, a phenomenon that gives rise to diverse non-membrane-bound nuclear, cytoplasmic and extracellular compartments4. We show that Drosophila HP1a protein undergoes liquid–liquid demixing in vitro, and nucleates into foci that display liquid properties during the first stages of heterochromatin domain formation in early Drosophila embryos. Furthermore, in both Drosophila and mammalian cells, heterochromatin domains exhibit dynamics that are characteristic of liquid phase-separation, including sensitivity to the disruption of weak hydrophobic interactions, and reduced diffusion, increased coordinated movement and inert probe exclusion at the domain boundary. We conclude that heterochromatic domains form via phase separation, and mature into a structure that includes liquid and stable compartments. We propose that emergent biophysical properties associated with phase-separated systems are critical to understanding the unusual behaviours of heterochromatin, and how chromatin domains in general regulate essential nuclear functions.

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supporting

confidence: 91%

Phase separation drives heterochromatin domain formation

Strom

Emelyanov

Mir

et al. 2017

Nature

1,639

1,431

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“…In somatic cells, NPM1 − one of the NPM2 homologs − undergoes phase separation in a concentration-dependent manner together with the arginine-rich proteins and rRNAs that are required for ribosome biogenesis. This event is critical for structural organization of the somatic nucleoli and, presumably, required to control the activity of ribosome biogenesis (Feric et al, 2016;Holmberg Olausson et al, 2014;Mitrea et al, 2016). In our study, we found that NPM2 has the ability to form a spherical body together with other proteins and RNAs in a concentration-dependent manner.…”

Section: (N>3 Each) (B) Npm2 Protein Expression Levels Insupporting

confidence: 48%

“…Because of the complexity of biochemical experiments when using mammalian oocytes, there are few biochemical data of mammalian NPM2 to specify its molecular function. However, one can speculate about the function of NPM2 by comparing it to NPM1 in somatic cells and to nucleoplasmin-like protein (NPL) in Drosophila, since both proteins have the conserved region of the NPM-core domain and also form a spherical body (Liu et al, 2006;Mitrea et al, 2016). For example, NPM1, has the ability to bind to the centromere-specific nucleosome CENP-A and is required for proper kinetochore and microtubule attachments (Amin et al, 2008;Dunleavy et al, 2009;Foltz et al, 2009).…”

Section: Discussionmentioning

confidence: 99%

Reconstitution of the oocyte nucleolus in mice through a single nucleolar protein, NPM2

Ogushi

Yamagata²,

Obuse

et al. 2017

Journal of Cell Science

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The mammalian oocyte nucleolus, the most prominent subcellular organelle in the oocyte, is vital in early development, yet its key functions and constituents remain unclear. We show here that the parthenotes/zygotes derived from enucleolated oocytes exhibited abnormal heterochromatin formation around parental pericentromeric DNAs, which led to a significant mitotic delay and frequent chromosome mis-segregation upon the first mitotic division. A proteomic analysis identified nucleoplasmin 2 (NPM2) as a dominant component of the oocyte nucleolus. Consistently, Npm2-deficient oocytes, which lack a normal nucleolar structure, showed chromosome segregation defects similar to those in enucleolated oocytes, suggesting that nucleolar loss, rather than micromanipulation-related damage to the genome, leads to a disorganization of higher-order chromatin structure in pronuclei and frequent chromosome mis-segregation during the first mitosis. Strikingly, expression of NPM2 alone sufficed to reconstitute the nucleolar structure in enucleolated embryos, and rescued their first mitotic division and full-term development. The nucleolus rescue through NPM2 required the pentamer formation and both the N-and C-terminal domains. Our findings demonstrate that the NPM2-based oocyte nucleolus is an essential platform for parental chromatin organization in early embryonic development.

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“…As a histone chaperone, NPM1 binds most efficiently to acetylated H3/H4 heterodimers and, as mentioned above, is able to promote the activities of nucleosome remodeling complexes and to support transcription and chromatin packaging activities (Okuwaki et al 2001;Swaminathan et al 2005). The ability of NPM1 to interact with highly positive sequences, along with its oligomerization activity have D r a f t 9 led several groups to propose that NPM1 may be the scaffolding protein responsible for retention of proteins containing arginine/lysine-rich nucleolar localization signals (NoLSs) within the nucleolus (Emmott and Hiscox 2009;Li et al 1996;Mitrea et al 2016). Through its C-terminal regions, NPM1 is able to bind short ssDNA sequences and promote dsDNA strand annealing (Borggrefe et al 1998) and, independently of those functions, to also interact with structured RNAs in response to genotoxic stress (Yang et al 2002).…”

Section: Nucleophosmin and Nucleolin: Two Multifunctional Nucleolar Pmentioning

confidence: 99%

Nucleolin and nucleophosmin: nucleolar proteins with multiple functions in DNA repair

Scott

Oeffinger

2016

Biochem. Cell Biol.

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The nucleolus represents a highly multifunctional intranuclear organelle in which, in addition to the canonical ribosome assembly, numerous processes such as transcription, DNA repair and replication, the cell cycle and apoptosis are coordinated. The nucleolus is further a key hub in the sensing of cellular stress and undergoes major structural and compositional changes in response to cellular perturbations.Numerous nucleolar proteins have been identified that, upon nucleolar stress sensing, deploy additional, non-ribosomal roles in the regulation of varied cell processes including cell cycle arrest, arrest of DNA replication, induction of DNA repair, and apoptosis, among others. The highly abundant proteins nucleophosmin (NPM1) and nucleolin (NCL) are two such factors that transit to the nucleoplasm in response to stress, and participate directly in the repair of numerous different DNA damages. This review discusses the contributions made by NCL and/or NPM1 to the different DNA repair pathways employed by mammalian cells to repair DNA insults, and examines the implications of such activities for the regulation, pathogenesis and therapeutic targeting of NPM1 and NCL.

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Nucleophosmin integrates within the nucleolus via multi-modal interactions with proteins displaying R-rich linear motifs and rRNA

Cited by 446 publications

References 70 publications

Phase separation drives heterochromatin domain formation

Phase separation drives heterochromatin domain formation

Reconstitution of the oocyte nucleolus in mice through a single nucleolar protein, NPM2

Nucleolin and nucleophosmin: nucleolar proteins with multiple functions in DNA repair

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