Compromised paraspeckle formation as a pathogenic factor in FUSopathies

Shelkovnikova, Tatyana A.; Robinson, Hannah; Troakes, Claire; Ninkina, Natalia; Buchman, Vladimir L.

doi:10.1093/hmg/ddt622

Cited by 115 publications

(123 citation statements)

References 55 publications

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“…2B and Fig. S3), which is consistent with a previous report (19). With the N-terminal 1-164 deletion, the punctate pattern disappeared and the FUS protein was evenly distributed in the entire nucleus including nucleoli.…”

Section: The N-terminal Qgsy-rich Region Is Responsible For Fus Chromsupporting

confidence: 92%

Self-assembled FUS binds active chromatin and regulates gene transcription

Yang

Gál

Chen

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

126

122

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Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease. Fused in sarcoma (FUS) is a DNA/RNA binding protein and mutations in FUS cause a subset of familial ALS. Most ALS mutations are clustered in the C-terminal nuclear localization sequence of FUS and consequently lead to the accumulation of protein inclusions in the cytoplasm. It remains debatable whether loss of FUS normal function in the nucleus or gain of toxic function in the cytoplasm plays a more critical role in the ALS etiology. Moreover, the physiological function of FUS in the nucleus remains to be fully understood. In this study, we found that a significant portion of nuclear FUS was bound to active chromatin and that the ALS mutations dramatically decreased FUS chromatin binding ability. Functionally, the chromatin binding is required for FUS transcription activation, but not for alternative splicing regulation. The N-terminal QGSY (glutamine-glycine-serine-tyrosine)-rich region (amino acids 1-164) mediates FUS self-assembly in the nucleus of mammalian cells and the self-assembly is essential for its chromatin binding and transcription activation. In addition, RNA binding is also required for FUS self-assembly and chromatin binding. Together, our results suggest a functional assembly of FUS in the nucleus under physiological conditions, which is different from the cytoplasmic inclusions. The ALS mutations can cause loss of function in the nucleus by disrupting this assembly and chromatin binding. FUS is a multifunctional protein and has been reported to play a role in various aspects of RNA metabolism (6), including transcription regulation and alternative splicing. FUS was initially identified in liposarcomas as part of a fusion protein (7,8) in which the N-terminal domain of FUS (amino acid 1-266) is recombined to transcription factor CHOP at its N terminus. The FUS-CHOP fusion protein activates the transcription of oncogenes and promotes tumorigenesis (9, 10). In familial ALS, most mutations are clustered in the C-terminal nuclear localization sequence (NLS) of FUS and consequently cause the mislocalization of FUS protein from the nucleus to the cytoplasm and the accumulation of protein inclusions (11-13). Such observations suggest two potential disease-causing mechanisms: loss of FUS normal function in the nucleus and gain of toxic function in the cytoplasm. It remains to be determined which mechanism plays a more critical role in ALS etiology and the two mechanisms are not necessarily exclusive of each other.Cytoplasmic FUS inclusions resemble stress granules, indicated by colocalization of FUS with different stress granule components (11,12). Stress granules are temporary cellular structures containing RNAs and proteins from suspended translation apparatus (14). Stress granule formation promotes cell survival under stressed conditions by redistributing translation resources. Compromised stress granule response in the presence of FUS mutants is considered a contributing factor to motor neuron dysfunction (15).Although t...

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Section: The N-terminal Qgsy-rich Region Is Responsible For Fus Chromsupporting

confidence: 92%

Self-assembled FUS binds active chromatin and regulates gene transcription

Yang

Gál

Chen

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

126

122

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“…For example, proteomic analysis of TDP-43 aggregates showed deposition of stress granule proteins G3BP and PABPC1 as well as paraspeckle proteins PSF and NONO (Dammer et al, 2012). Similar observations of paraspeckle proteins p54nrb and NONO in FUS-positive inclusions (Shelkovnikova et al, 2014) provide additional evidence in support of this concept. Thus, understanding the protein-protein interactions (PPIs) of ALS-linked RBPs is a necessary step towards defining the protein composition of inclusions in ALS/FTLD and new insight into mechanisms of disease.…”

Section: Introductionsupporting

confidence: 72%

Immunoprecipitation and mass spectrometry defines an extensive RBM45 protein–protein interaction network

Collins

et al. 2016

Brain Research

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The pathological accumulation of RNA-binding proteins (RBPs) within inclusion bodies is a hallmark of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). RBP aggregation results in both toxic gain and loss of normal function. Determining the protein binding partners and normal functions of disease-associated RBPs is necessary to fully understand molecular mechanisms of RBPs in disease. Herein, we characterized the protein-protein interactions (PPIs) of RBM45, a RBP that localizes to inclusions in ALS/FTLD. Using immunoprecipitation coupled to mass spectrometry (IP-MS), we identified 132 proteins that specifically interact with RBM45 within HEK293 cells. Select PPIs were validated by immunoblot and immunocytochemistry, demonstrating that RBM45 associates with a number of other RBPs primarily via RNA-dependent interactions in the nucleus. Analysis of the biological processes and pathways associated with RBM45-interacting proteins indicates enrichment for nuclear RNA processing/splicing via association with hnRNP proteins and cytoplasmic RNA translation via eiF2 and eiF4 pathways. Moreover, several other ALS-linked RBPs, including TDP-43, FUS, Matrin-3, and hnRNP-A1, interact with RBM45, consistent with prior observations of these proteins within intracellular inclusions in ALS/FTLD. Taken together, our results define a PPI network for RBM45, suggest novel functions for this protein, and provide new insights into the contributions of RBM45 to neurodegeneration in ALS/FTLD.

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“…It is possible that MCF7 lacks the expression of essential factors activating c-Myc translation. This hypothesis is supported by the observation that although P54nrb and PSF are expressed in MCF7, localization of P54nrb and PSF is predominantly in the perinucleolar regions instead of paraspeckles in some cells [27]. MCF7 lacks enzymes ( e .…”

Section: Discussionmentioning

confidence: 84%

Depletion of NEAT1 lncRNA attenuates nucleolar stress by releasing sequestered P54nrb and PSF to facilitate c-Myc translation

et al. 2017

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Altered expression of NEAT1, the architectural long non-coding RNA (lncRNA) of nuclear paraspeckles, has been reported during tumorigenesis, as well as under various cellular stress conditions. Here we report that the depletion of NEAT1 lncRNA alleviates nucleolar stress during RNAP I inhibition through releasing sequestered P54nrb and PSF to facilitate the IRES-dependent translation of c-Myc. RNAP I inhibitor CX5461 disrupts the SL1-rDNA interaction and induces nucleolar disruption, demonstrated by the accumulation of fibrillarin-containing nucleoplasmic foci and nucleolar clearance of ribosomal proteins in HeLa cells. Antisense oligonucleotide-mediated depletion of NEAT1 lncRNA significantly attenuated the RNAP I inhibition and its related nucleolar disruption. Interestingly, induction in the levels of c-Myc protein was observed in NEAT1-depeleted cells under RNAP I inhibition. NEAT1-associated paraspeckle proteins P54nrb and PSF have been reported as positive regulators of c-Myc translation through interaction with c-Myc IRES. Indeed, an increased association of P54nrb and PSF with c-Myc mRNA was observed in NEAT1-depleted cells. Moreover, apoptosis was observed in HeLa cells depleted of P54nrb and PSF, further confirming the positive involvement of P54nrb and PSF in cell proliferation. Together, our results suggest that NEAT1 depletion rescues CX5461-induced nucleolar stress through facilitating c-Myc translation by relocating P54nrb/PSF from nuclear paraspeckles to c-Myc mRNAs.

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Compromised paraspeckle formation as a pathogenic factor in FUSopathies

Cited by 115 publications

References 55 publications

Self-assembled FUS binds active chromatin and regulates gene transcription

Self-assembled FUS binds active chromatin and regulates gene transcription

Immunoprecipitation and mass spectrometry defines an extensive RBM45 protein–protein interaction network

Depletion of NEAT1 lncRNA attenuates nucleolar stress by releasing sequestered P54nrb and PSF to facilitate c-Myc translation

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