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The pathological accumulation of the microtubule binding protein tau drives age-related neurodegeneration in a variety of disorders, collectively called tauopathies. In the most common tauopathy, Alzheimer's disease (AD), the accumulation of pathological tau strongly correlates with cognitive decline. The underlying molecular mechanisms that drive neurodegeneration in tauopathies remain incompletely understood and no effective disease modifying pharmacological interventions currently exist. Here, we show that tau toxicity depends on the highly conserved nuclear E3 ubiquitin ligase adaptor protein SPOP in a Caenorhabditis elegans model of tauopathy. Loss of function mutations in the C. elegans spop-1 gene significantly improves behavioral deficits in tau transgenic animals, while neuronal overexpression of SPOP-1 protein significantly worsens behavioral deficits. In addition, loss of spop-1 rescues a variety of tau-related phenotypes including the accumulation of total and phosphorylated tau protein, neurodegeneration, and shortened lifespan. Knockdown of SPOP-1's E3 ubiquitin ligase cul-3/Cullin3 does not improve tauopathy suggesting a non-degradative mechanism of action for SPOP-1. Suppression of disease-related phenotypes occurs independently of the nuclear speckle resident poly(A)-binding protein SUT-2/MSUT2. MSUT2 modifies tauopathy in mammalian neurons and in AD. Our work identifies SPOP as a novel modifier of tauopathy and a conceptual pathway for therapeutic intervention. [ABSTRACT FROM AUTHOR]

A site-directed mutagenesis study was conducted to explore the significance of residues Ser175, phenylalanine Phe115, and threonine Thr178 in Escherichia coli towards the oxidation of dihydroorotate (DHO). Changing the size and/or hydrophobicity of the residues of the hydrogen bond network, Thr178 and Phe115 which slowed flavin reduction as much as 2 orders of magnitude, indicated that the active site base and the hydrogen bond network work together for efficient deprotonation of DHO.

G-protein-coupled receptors (GPCRs) are the darling drug targets of many pharmaceutical and biotech companies. This largest superfamily of cell-membrane receptors affects many aspects of life, including mood and behaviour, the [...]

One of the major signs of severe organophosphate poisoning is seizures. Previous studies have shown that both muscarinic agonist- and organophosphate-induced seizures require activation of muscarinic acetylcholine receptors in the central nervous system. Seizures induced by the muscarinic agonist pilocarpine require the M1 receptor and are modulated by cannabinoid CB1 receptors. In this study, we determined whether M1 and CB1 receptors also regulated seizures induced by the organophosphate paraoxon. We found no differences in seizures induced by paraoxon in wild-type ( WT) and M1 knockout ( KO) mice, indicating that in contrast to pilocarpine seizures, M1 receptors are not required for paraoxon seizures. Furthermore, we found that pilocarpine administration resulted in seizure-independent activation of ERK in the hippocampus in a M1 receptor-dependent manner, while paraoxon did not induce seizure-independent activation of ERK in the mouse hippocampus. This shows that pilocarpine and paraoxon activated M1 receptors in the hippocampus to different extents. There were no differences in seizures induced by paraoxon in WT and CB1 KO mice, and neither CB1 agonist nor antagonist administration had significant effects on paraoxon seizures, indicating that, in contrast to pilocarpine seizures, paraoxon seizures are not modulated by CB1 receptors. These results demonstrate that there are fundamental molecular differences in the regulation of seizures induced by pilocarpine and paraoxon. [ABSTRACT FROM AUTHOR]

Receptor internalization from the cell surface occurs through several mechanisms. Some of these mechanisms, such as clathrin coated pits, are well understood. The M2 muscarinic acetylcholine receptor undergoes internalization via a poorlydefined clathrin-independent mechanism. We used isotope coded affinity tagging and mass spectrometry to identify the scaffolding protein, receptor for activated C kinase (RACK1) as a protein enriched in M2-immunoprecipitates from M2-expressing cells over those of non-M2 expressing cells. Treatment of cells with the agonist carbachol disrupted the interaction of RACK1 with M2. We further found that RACK1 overexpression inhibits the internalization and subsequent down regulation of the M2 receptor in a receptor subtype-specific manner. Decreased RACK1 expression increases the rate of agonist internalization of the M2 receptor, but decreases the extent of subsequent down-regulation. These results suggest that RACK1 may both interfere with agonist-induced sequestration and be required for subsequent targeting of internalized M2 receptors to the degradative pathway. [ABSTRACT FROM AUTHOR]

Genetic variation in the lysosomal and transmembrane protein 106B (TMEM106B) modifies risk for a diverse range of neurodegenerative disorders, especially frontotemporal lobar degeneration (FTLD) with progranulin (PGRN) haplo-insufficiency, although the molecular mechanisms involved are not yet understood. Through advances in cryo-electron microscopy (cryo-EM), homotypic aggregates of the C-Terminal domain of TMEM106B (TMEM CT) were discovered as a previously unidentified cytosolic proteinopathy in the brains of FTLD, Alzheimer's disease, progressive supranuclear palsy (PSP), and dementia with Lewy bodies (DLB) patients. While it remains unknown what role TMEM CT aggregation plays in neuronal loss, its presence across a range of aging related dementia disorders indicates involvement in multi-proteinopathy driven neurodegeneration. To determine the TMEM CT aggregation propensity and neurodegenerative potential, we characterized a novel transgenic C. elegans model expressing the human TMEM CT fragment constituting the fibrillar core seen in FTLD cases. We found that pan-neuronal expression of human TMEM CT in C. elegans causes neuronal dysfunction as evidenced by behavioral analysis. Cytosolic aggregation of TMEM CT proteins accompanied the behavioral dysfunction driving neurodegeneration, as illustrated by loss of GABAergic neurons. To investigate the molecular mechanisms driving TMEM106B proteinopathy, we explored the impact of PGRN loss on the neurodegenerative effect of TMEM CT expression. To this end, we generated TMEM CT expressing C. elegans with loss of pgrn-1 , the C. elegans ortholog of human PGRN. Neither full nor partial loss of pgrn-1 altered the motor phenotype of our TMEM CT model suggesting TMEM CT aggregation occurs downstream of PGRN loss of function. We also tested the ability of genetic suppressors of tauopathy to rescue TMEM CT pathology. We found that genetic knockout of spop-1, sut-2, and sut-6 resulted in weak to no rescue of proteinopathy phenotypes, indicating that the mechanistic drivers of TMEM106B proteinopathy may be distinct from tauopathy. Taken together, our data demonstrate that TMEM CT aggregation can kill neurons. Further, expression of TMEM CT in C. elegans neurons provides a useful model for the functional characterization of TMEM106B proteinopathy in neurodegenerative disease., Competing Interests: Competing interests The authors have no competing interests to declare.

One of the major signs of severe organophosphate poisoning is seizures. Previous studies have shown that both muscarinic agonist- and organophosphate-induced seizures require activation of muscarinic acetylcholine receptors in the central nervous system. Seizures induced by the muscarinic agonist pilocarpine require the M1 receptor and are modulated by cannabinoid CB1 receptors. In this study, we determined whether M1 and CB1 receptors also regulated seizures induced by the organophosphate paraoxon. We found no differences in seizures induced by paraoxon in wild-type (WT) and M1 knockout (KO) mice, indicating that in contrast to pilocarpine seizures, M1 receptors are not required for paraoxon seizures. Furthermore, we found that pilocarpine administration resulted in seizure-independent activation of ERK in the hippocampus in a M1 receptor-dependent manner, while paraoxon did not induce seizure-independent activation of ERK in the mouse hippocampus. This shows that pilocarpine and paraoxon activated M1 receptors in the hippocampus to different extents. There were no differences in seizures induced by paraoxon in WT and CB1 KO mice, and neither CB1 agonist nor antagonist administration had significant effects on paraoxon seizures, indicating that, in contrast to pilocarpine seizures, paraoxon seizures are not modulated by CB1 receptors. These results demonstrate that there are fundamental molecular differences in the regulation of seizures induced by pilocarpine and paraoxon.