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The Journal of Biological Chemistry Aug 2023Paraspeckles (PS) are nuclear structures scaffolded by the long noncoding RNA NEAT1 and protein components such as NONO and SFPQ. We previously found that the...
Paraspeckles (PS) are nuclear structures scaffolded by the long noncoding RNA NEAT1 and protein components such as NONO and SFPQ. We previously found that the upregulation of RNA N6-methyl-adenosine (mA) demethylase ALKBH5 facilitates hypoxia-induced paraspeckle assembly through erasing mA marks on NEAT1, thus stabilizing it. However, it remains unclear how these processes are spatiotemporally coordinated. Here we discover that ALKBH5 specifically binds to proteins in PS and forms phase-separated droplets that are incorporated into PS through its C-terminal intrinsically disordered region (cIDR). Upon exposure to hypoxia, rapid ALKBH5 condensation in PS induces mA demethylation of NEAT1, which further facilitates PS formation before the upregulation of ALKBH5 expression. In cells expressing ALKBH5 lacking cIDR, PS fail to be formed in response to hypoxia, accompanied with insufficient mA demethylation of NEAT1 and its destabilization. We also demonstrate that ALKBH5-cIDR is indispensable for hypoxia-induced effects such as cancer cell invasion. Therefore, our study has identified the role of ALKBH5 in phase separation as the molecular basis of the positive feedback loop for PS formation between ALKBH5 incorporation into PS and NEAT1 stabilization.
Topics: Humans; AlkB Homolog 5, RNA Demethylase; Hypoxia; Paraspeckles; RNA, Long Noncoding; Transcriptional Activation; Up-Regulation
PubMed: 37474102
DOI: 10.1016/j.jbc.2023.105071 -
Cell Cycle (Georgetown, Tex.) Jan 2017
Topics: DNA Damage; Humans; Intranuclear Inclusion Bodies; Models, Biological; Neoplasms; RNA, Long Noncoding; Stress, Physiological
PubMed: 27700225
DOI: 10.1080/15384101.2016.1235847 -
The EMBO Journal Jun 2021Paraspeckles are constructed by NEAT1_2 architectural long noncoding RNAs. Their characteristic cylindrical shapes, with highly ordered internal organization,...
Paraspeckles are constructed by NEAT1_2 architectural long noncoding RNAs. Their characteristic cylindrical shapes, with highly ordered internal organization, distinguish them from typical liquid-liquid phase-separated condensates. We experimentally and theoretically investigated how the shape and organization of paraspeckles are determined. We identified the NEAT1_2 RNA domains responsible for shell localization of the NEAT1_2 ends, which determine the characteristic internal organization. Using the soft matter physics, we then applied a theoretical framework to understand the principles that determine NEAT1_2 organization as well as shape, number, and size of paraspeckles. By treating paraspeckles as amphipathic block copolymer micelles, we could explain and predict the experimentally observed behaviors of paraspeckles upon NEAT1_2 domain deletions or transcriptional modulation. Thus, we propose that paraspeckles are block copolymer micelles assembled through a type of microphase separation, micellization. This work provides an experiment-based theoretical framework for the concept that ribonucleoprotein complexes (RNPs) can act as block copolymers to form RNA-scaffolding biomolecular condensates with optimal sizes and structures in cells.
Topics: Cell Line; Humans; Micelles; Polymers; RNA, Long Noncoding; Ribonucleoproteins
PubMed: 33885174
DOI: 10.15252/embj.2020107270 -
Stem Cell Reports Dec 2020Membrane-free intracellular biocondensates are enclosures of proteins and nucleic acids that form by phase separation. Extensive ensembles of nuclear "membraneless... (Review)
Review
Membrane-free intracellular biocondensates are enclosures of proteins and nucleic acids that form by phase separation. Extensive ensembles of nuclear "membraneless organelles" indicate their involvement in genome regulation. Indeed, nuclear bodies have been linked to regulation of gene expression by formation of condensates made of chromatin and RNA processing factors. Important questions pertain to the involvement of membraneless organelles in determining cell identity through their cell-type-specific composition and function. Paraspeckles provide a prism to these questions because they exhibit striking cell-type-specific patterns and since they are crucial in embryogenesis. Here, we outline known interactions between paraspeckles and chromatin, and postulate how such interactions may be important in regulation of cell fate transitions. Moreover, we propose long non-coding RNAs (lncRNAs) as candidates for similar regulation because many form foci that resemble biocondensates and exhibit dynamic patterns during differentiation. Finally, we outline approaches that could ascertain how chromatin-associated membraneless organelles regulate cellular differentiation.
Topics: Animals; Cell Differentiation; Chromatin; Embryo, Mammalian; Embryonic Development; Humans; Organelles; RNA Processing, Post-Transcriptional; RNA, Long Noncoding
PubMed: 33217325
DOI: 10.1016/j.stemcr.2020.10.011 -
Frontiers in Molecular Biosciences 2021RNA molecules are increasingly being identified as facilitating or impeding the interaction of proteins and nucleic acids, serving as so-called scaffolds or decoys. Long... (Review)
Review
RNA molecules are increasingly being identified as facilitating or impeding the interaction of proteins and nucleic acids, serving as so-called scaffolds or decoys. Long non-coding RNAs have been commonly implicated in such roles, particularly in the regulation of nuclear processes including chromosome topology, regulation of chromatin state and gene transcription, and assembly of nuclear biomolecular condensates such as paraspeckles. Recently, an increased awareness of cytoplasmic RNA scaffolds and decoys has begun to emerge, including the identification of non-coding regions of mRNAs that can also function in a scaffold-like manner to regulate interactions of nascently translated proteins. Collectively, cytoplasmic RNA scaffolds and decoys are now implicated in processes such as mRNA translation, decay, protein localization, protein degradation and assembly of cytoplasmic biomolecular condensates such as P-bodies. Here, we review examples of RNA scaffolds and decoys in both the nucleus and cytoplasm, illustrating common themes, the suitability of RNA to such roles, and future challenges in identifying and better understanding RNA scaffolding and decoy functions.
PubMed: 33898516
DOI: 10.3389/fmolb.2021.634146 -
Frontiers in Molecular Biosciences 2022Phase separation is a fundamental mechanism for compartmentalization in cells and leads to the formation of biomolecular condensates, generally containing various RNA... (Review)
Review
Phase separation is a fundamental mechanism for compartmentalization in cells and leads to the formation of biomolecular condensates, generally containing various RNA molecules. RNAs are biomolecules that can serve as suitable scaffolds for biomolecular condensates and determine their forms and functions. Many studies have focused on biomolecular condensates formed by liquid-liquid phase separation (LLPS), one type of intracellular phase separation mechanism. We recently identified that paraspeckle nuclear bodies use an intracellular phase separation mechanism called micellization of block copolymers in their formation. The paraspeckles are scaffolded by NEAT1_2 long non-coding RNAs (lncRNAs) and their partner RNA-binding proteins (NEAT1_2 RNA-protein complexes [RNPs]). The NEAT1_2 RNPs act as block copolymers and the paraspeckles assemble through micellization. In LLPS, condensates grow without bound as long as components are available and typically have spherical shapes to minimize surface tension. In contrast, the size, shape, and internal morphology of the condensates are more strictly controlled in micellization. Here, we discuss the potential importance and future perspectives of micellization of block copolymers of RNPs in cells, including the construction of designer condensates with optimal internal organization, shape, and size according to design guidelines of block copolymers.
PubMed: 36106018
DOI: 10.3389/fmolb.2022.974772 -
Frontiers in Cellular Neuroscience 2022Understanding and ameliorating neurodegenerative diseases represents a key challenge for supporting the health span of the aging population. Diverse protein aggregates... (Review)
Review
Understanding and ameliorating neurodegenerative diseases represents a key challenge for supporting the health span of the aging population. Diverse protein aggregates have been implicated in such neurodegenerative disorders, including amyloid-β, α-synuclein, tau, fused in sarcoma (FUS), and transactivation response element (TAR) DNA-binding protein 43 (TDP-43). Recent years have seen significant growth in our mechanistic knowledge of relationships between these proteins and some of the membrane-less nuclear structures that fulfill key roles in the cell function. These include the nucleolus, nuclear speckles, and paraspeckles. The ability of macromolecular protein:RNA complexes to partition these nuclear condensates through biophysical processes that involve liquid-liquid phase separation (LLPS) has also gained attention recently. The paraspeckle, which is scaffolded by the architectural long-non-coding RNA nuclear enriched abundant transcript 1 (NEAT1) plays central roles in RNA processing and metabolism and has been linked dynamically to TDP-43. In this mini-review, we outline essential early and recent insights in relation to TDP-43 proteinopathies. We then appraise the relationships between TDP-43 and NEAT1 in the context of neuronal paraspeckles and neuronal stress. We highlight key areas for investigation based on recent advances in our understanding of how TDP-43 affects neuronal function, especially in relation to messenger ribosomal nucleic acid (mRNA) splicing. Finally, we offer perspectives that should be considered for translational pipelines in order to improve health outcomes for the management of neurodegenerative diseases.
PubMed: 36385948
DOI: 10.3389/fncel.2022.954912 -
Cellular and Molecular Life Sciences :... Oct 2020Nuclear paraspeckle assembly transcript 1 (NEAT1) is a long non-coding RNA that is widely expressed in a variety of mammalian cell types. An increasing number of studies... (Review)
Review
Nuclear paraspeckle assembly transcript 1 (NEAT1) is a long non-coding RNA that is widely expressed in a variety of mammalian cell types. An increasing number of studies have demonstrated that NEAT1 plays key roles in various biological and pathological processes; therefore, it is important to understand how its expression is regulated and how it regulates the expression of its target genes. Recently, we found that NEAT1 expression could be regulated by signal transducer and activator of transcription 3 and that altered NEAT1 expression epigenetically regulates downstream gene transcription during herpes simplex virus-1 infection and Alzheimer's disease, suggesting that NEAT1 acts as an important sensor and effector during stress and disease development. In this review, we summarize and discuss the molecules and regulatory patterns that control NEAT1 gene expression and the molecular mechanism via which NEAT1 regulates the expression of its target genes, providing novel insights into the central role of NEAT1 in gene regulation.
Topics: Epithelial-Mesenchymal Transition; Gene Expression Regulation; Humans; MicroRNAs; RNA 3' End Processing; RNA Splicing; RNA Stability; RNA, Long Noncoding; RNA-Binding Proteins; Transcription Factors
PubMed: 32219465
DOI: 10.1007/s00018-020-03503-0 -
The EMBO Journal Nov 2022Several kinds of stress promote the formation of three-stranded RNA:DNA hybrids called R-loops. Insufficient clearance of these structures promotes genomic instability...
Several kinds of stress promote the formation of three-stranded RNA:DNA hybrids called R-loops. Insufficient clearance of these structures promotes genomic instability and DNA damage, which ultimately contribute to the establishment of cancer phenotypes. Paraspeckle assemblies participate in R-loop resolution and preserve genome stability, however, the main determinants of this mechanism are still unknown. This study finds that in Multiple Myeloma (MM), AATF/Che-1 (Che-1), an RNA-binding protein fundamental to transcription regulation, interacts with paraspeckles via the lncRNA NEAT1_2 (NEAT1) and directly localizes on R-loops. We systematically show that depletion of Che-1 produces a marked accumulation of RNA:DNA hybrids. We provide evidence that such failure to resolve R-loops causes sustained activation of a systemic inflammatory response characterized by an interferon (IFN) gene expression signature. Furthermore, elevated levels of R-loops and of mRNA for paraspeckle genes in patient cells are linearly correlated with Multiple Myeloma progression. Moreover, increased interferon gene expression signature in patients is associated with markedly poor prognosis. Taken together, our study indicates that Che-1/NEAT1 cooperation prevents excessive inflammatory signaling in Multiple Myeloma by facilitating the clearance of R-loops. Further studies on different cancer types are needed to test if this mechanism is ubiquitously conserved and fundamental for cell homeostasis.
Topics: Humans; R-Loop Structures; Multiple Myeloma; Paraspeckles; RNA, Long Noncoding; Interferons; Repressor Proteins; Apoptosis Regulatory Proteins
PubMed: 35929179
DOI: 10.15252/embj.2021109711 -
Nucleus (Austin, Tex.) May 2017Circadian clocks regulate rhythmic gene expression levels by means of mRNA oscillations that are mainly driven by post-transcriptional regulation. We identified a new... (Review)
Review
Circadian clocks regulate rhythmic gene expression levels by means of mRNA oscillations that are mainly driven by post-transcriptional regulation. We identified a new post-transcriptional mechanism, which involves nuclear bodies called paraspeckles. Major components of paraspeckles including the long noncoding RNA Neat1, which is the structural component, and its major protein partners, as well as the number of paraspeckles, follow a circadian pattern in pituitary cells. Paraspeckles are known to retain within the nucleus RNAs containing inverted repeats of Alu sequences. We showed that a reporter gene in which these RNA duplex elements were inserted in the 3'-UTR region displayed a circadian expression. Moreover, circadian endogenous mRNA associated with paraspeckles lost their circadian pattern when paraspeckles were disrupted. This work not only highlights a new paraspeckle-based post-transcriptional mechanism involved in circadian gene expression but also provides the list of all mRNA associated with paraspeckles in the nucleus of pituitary cells.
Topics: Animals; Cell Nucleus; Circadian Rhythm; Gene Expression Regulation; Pituitary Gland; RNA, Messenger
PubMed: 28060565
DOI: 10.1080/19491034.2016.1277304