Pur-alpha regulates cytoplasmic stress granule dynamics and ameliorates FUS toxicity

Daigle, J. Gavin; Krishnamurthy, Karthik; Ramesh, Nandini; Casci, Ian; Monaghan, John M.; McAvoy, Kevin; Godfrey, Earl W.; Daniel, Dianne C.; Johnson, Edward M.; Monahan, Zachary; Shewmaker, Frank; Pasinelli, Piera; Pandey, Udai Bhan

doi:10.1007/s00401-015-1530-0

Cited by 61 publications

(77 citation statements)

References 75 publications

(105 reference statements)

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“…Recent reports suggest these granules are membraneless organelles, partially or wholly stabilized by liquid–liquid phase separation (LLPS) where molecules self segregate (demix) to form two co‐existing liquids as observed for oil droplets in water (Courchaine et al , 2016). However, unlike oil–water emulsions, RNP granules formed by LLPS are not devoid of water and are thought to be stabilized by multivalent, dynamic protein:protein and protein:mRNA interactions, concentrating granule components into a dense and viscous compartment into which other molecules can partition and localizing their cellular function (Ryu et al , 2014; Burke et al , 2015; Di Salvio et al , 2015; Lenzi et al , 2015; Bowden & Dormann, 2016; Daigle et al , 2016; Nott et al , 2016). Indeed, FUS‐containing granules and FUS at sites of DNA damage have liquid‐like properties in cells (Patel et al , 2015).…”

Section: Introductionmentioning

confidence: 99%

Phosphorylation of the FUS low‐complexity domain disrupts phase separation, aggregation, and toxicity

et al. 2017

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Neuronal inclusions of aggregated RNA‐binding protein fused in sarcoma (FUS) are hallmarks of ALS and frontotemporal dementia subtypes. Intriguingly, FUS's nearly uncharged, aggregation‐prone, yeast prion‐like, low sequence‐complexity domain (LC) is known to be targeted for phosphorylation. Here we map in vitro and in‐cell phosphorylation sites across FUS LC. We show that both phosphorylation and phosphomimetic variants reduce its aggregation‐prone/prion‐like character, disrupting FUS phase separation in the presence of RNA or salt and reducing FUS propensity to aggregate. Nuclear magnetic resonance spectroscopy demonstrates the intrinsically disordered structure of FUS LC is preserved after phosphorylation; however, transient domain collapse and self‐interaction are reduced by phosphomimetics. Moreover, we show that phosphomimetic FUS reduces aggregation in human and yeast cell models, and can ameliorate FUS‐associated cytotoxicity. Hence, post‐translational modification may be a mechanism by which cells control physiological assembly and prevent pathological protein aggregation, suggesting a potential treatment pathway amenable to pharmacologic modulation.

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Section: Introductionmentioning

confidence: 99%

Phosphorylation of the FUS low‐complexity domain disrupts phase separation, aggregation, and toxicity

et al. 2017

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“…Recent studies have also identified de novo mutations in the PURA gene, which encodes Pur-alpha, that are associated with neonatal hypotonia, encephalopathy with or without seizures, neurodevelopmental delays, learning disabilities, and poor coordination or ataxias in several young human subjects (Hunt et al, 2014, Lalani et al, 2014, Tanaka et al, 2015). Dysregulation of Pur-alpha has also been shown in C9Orf72-related Frontotemporal lobe dementia in association with Amyotrophic Lateral Sclerosis, presumably resulting from accumulation of intracellular inclusion bodies that contain Pur-alpha and RNA complexes, and RNA toxic gain-in-function effects (Daigle et al, 2016). Pur-alpha may even be involved in enhancing HIV-1 transcription, with increasing concentrations of Pur-alpha leading to increased transcription of HIV-1 (Chepenik et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

Memory deficits, gait ataxia and neuronal loss in the hippocampus and cerebellum in mice that are heterozygous for Pur-alpha

et al. 2016

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Pur-alpha is a highly conserved sequence specific DNA and RNA binding protein with established roles in DNA replication, RNA translation, cell cycle regulation, and maintenance of neuronal differentiation. Prior studies have shown that mice lacking Pur-alpha (−/−) display decreased neurogenesis and impaired neuronal differentiation. We sought to examine for the first time, the behavioral phenotype and brain histopathology of mice that are heterozygous (+/−) for Pur-alpha. Standardized behavioral phenotyping revealed a decreased escape response to touch, limb and abdominal hypotonia, and gait abnormalities in heterozygous Pur-alpha (+/−) mice, compared to wild-type (+/+) littermates. Footprint pattern analyses showed wider-based steps, increased missteps and more outwardly rotated hindpaws in heterozygous Pur-alpha (+/−) mice, suggestive of cerebellar pathology. Barnes maze and novel object location testing revealed significant memory deficits in heterozygous Pur-alpha mice, suggestive of hippocampal pathology. Quantitative immunohistochemical assays of the vermal region of the cerebellum and CA1-3 regions of the hippocampus revealed reduced numbers of neurons in general, as well as reduced numbers of Pur-alpha+-immunopositive neurons and dendrites in heterozygous Pur-alpha mice, compared to wild type littermates. Past studies have implicated mutations in Pur-alpha in several diseases of brain development and neurodegeneration. When combined with these new findings, the Pur-alpha heterozygous knockout mice may provide an animal model in which to study mechanisms of and treatments for Pur-alpha related cognitive deficiencies and neuropathology.

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“…) could further impair SG assembly in C9orf72 mutation carriers, as Pur‐α appears to be essential for SG formation (Daigle et al . ). Although these results have to be reconciled with the conflicting finding that TDP‐43 inclusions in C9orf72 patients frequently are SG marker (PABP1) positive (McGurk et al .…”

Section: Many Als/ftld‐associated Genes Affect Sg Dynamicsmentioning

confidence: 97%

Altered mRNP granule dynamics in FTLD pathogenesis

Bowden¹,

Dormann

2016

Journal of Neurochemistry

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In neurons, RNA-binding proteins (RBPs) play a key role in post-transcriptional gene regulation, for example alternative splicing, mRNA localization in neurites and local translation upon synaptic stimulation. There is increasing evidence that defective or mislocalized RBPs -and consequently altered mRNA processing -lead to neuronal dysfunction and cause neurodegeneration, including frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Cytosolic RBP aggregates containing TAR DNA-binding protein of 43 kDa (TDP-43) or fused in sarcoma (FUS) are a common hallmark of both disorders. There is mounting evidence that translationally silent mRNP granules, such as stress granules or transport granules, play an important role in the formation of these RBP aggregates. These granules are thought to be 'catalytic convertors' of RBP aggregation by providing a high local concentration of RBPs. As recently shown in vitro, RBPs that contain a so-called low-complexity domain start to 'solidify' and eventually aggregate at high protein concentrations. The same may happen in mRNP granules in vivo, leading to 'solidified' granules that lose their dynamic properties and ability to fulfill their physiological functions. This may result in a disturbed stress response, altered mRNA transport and local translation, and formation of pathological TDP-43 or FUS aggregates, all of which may contribute to neuronal dysfunction and neurodegeneration. Here, we discuss the general functional properties of these mRNP granules, how their dynamics may be disrupted in frontotemporal lobar degeneration/amyotrophic lateral sclerosis, for example by loss or gain of function of TDP-43 and FUS, and how this may contribute to the development of RBP aggregates and neurotoxicity.

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Pur-alpha regulates cytoplasmic stress granule dynamics and ameliorates FUS toxicity

Cited by 61 publications

References 75 publications

Phosphorylation of the FUS low‐complexity domain disrupts phase separation, aggregation, and toxicity

Phosphorylation of the FUS low‐complexity domain disrupts phase separation, aggregation, and toxicity

Memory deficits, gait ataxia and neuronal loss in the hippocampus and cerebellum in mice that are heterozygous for Pur-alpha

Altered mRNP granule dynamics in FTLD pathogenesis

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