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The correct subcellular distribution of proteins establishes the complex morphology and function of neurons. Fluorescence microscopy techniques are invaluable to investigate subcellular protein distribution, but they suffer from the limited ability to efficiently and reliably label endogenous proteins with fluorescent probes. We developed ORANGE: Open Resource for the Application of Neuronal Genome Editing, which mediates targeted genomic integration of epitope tags in rodent dissociated neuronal culture, in organotypic slices, and in vivo. ORANGE includes a knock-in library for in-depth investigation of endogenous protein distribution, viral vectors, and a detailed two-step cloning protocol to develop knock-ins for novel targets. Using ORANGE with (live-cell) superresolution microscopy, we revealed the dynamic nanoscale organization of endogenous neurotransmitter receptors and synaptic scaffolding proteins, as well as previously uncharacterized proteins. Finally, we developed a mechanism to create multiple knock-ins in neurons, mediating multiplex imaging of endogenous proteins. Thus, ORANGE enables quantification of expression, distribution, and dynamics for virtually any protein in neurons at nanoscale resolution. This study describes the development of a genome editing toolbox (ORANGE) for endogenous tagging of proteins in neurons. This open resource allows the investigation of protein localization and dynamics in neurons using live-cell and super-resolution imaging techniques. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is the most common cause of dementia worldwide with 10–30% prevalence in aging population and a high socioeconomic impact. Because AD definitive diagnostic requires post-mortem verification, new approaches to study the disease are necessary. Here, we analyze the humoral immune response in AD to survey whether APP+1 or UBB+1 frameshift proteins, produced as a consequence of the "molecular misreading" alteration in AD occurring in the APP (amyloid precursor protein) and UBB (ubiquitin-B protein) proteins' mRNA, elicit the production of autoantibodies specific of AD. To this end, APP+1 and UBB+1 peptides were expressed in bacteria as 6xHisHalo fusion proteins and after purification to homogeneity their seroreactivity was analyzed using 81 individual sera from AD patients and 43 individual sera from healthy individuals by luminescence beads immunoassay. We found that as a result of the molecular misreading, APP+1 and UBB+1 frameshift peptides produced a humoral immune response in AD patients, whose autoantibody levels are significantly higher in comparison with healthy controls. Their combination with a previously reported panel of four autoantigens specific of AD (ANTXR1, OR8J1, PYGB, and NUPR1) increased their diagnostic ability assessed by receiver operating characteristic (ROC) curves up to an area under the curve (AUC) of 73.5%. Collectively, our results demonstrate that APP+1 and UBB+1 frameshift proteins, non-previously described as AD-specific autoantigens, elicit the production of autoantibodies which might be useful as blood-based biomarkers to aid in the detection of the disease. [ABSTRACT FROM AUTHOR]

The ubiquitin-proteasome pathway (UPP) has multiple roles in the normal nervous system, including the development of synaptic connections and synaptic plasticity. Research over the past several years has indicated a role for the UPP in aging without any overt pathology in the brain. In addition, malfunction of the UPP is implicated in Alzheimer's disease (AD) and dementia associated with it. In this mini review article, we assess the literature on the role of protein degradation by the UPP in aging and in AD with special emphasis on dysregulation of the UPP and its contribution to cognitive decline and impairment. [ABSTRACT FROM AUTHOR]

Perineuronal nets (PNNs) are condensed structures in the extracellular matrix that mainly surround GABA-ergic parvalbumin-positive interneurons in the adult brain. Previous studies revealed a parallel between PNN formation and the closure of the critical period. Moreover, ocular dominance plasticity is enhanced in response to PNN manipulations in adult animals. However, the mechanisms through which perineuronal nets modulate plasticity are still poorly understood. Recent work indicated that perineuronal nets may convey molecular signals by binding and storing proteins with important roles in cellular communication. Here we report that semaphorin3A (Sema3A), a chemorepulsive axon guidance cue known to bind to important perineuronal net components, is necessary to dampen ocular dominance plasticity in adult rats. First, we showed that the accumulation of Sema3A in PNNs in the visual cortex correlates with critical period closure, following the same time course of perineuronal nets maturation. Second, the accumulation of Sema3A in perineuronal nets was significantly reduced by rearing animals in the dark in the absence of any visual experience. Finally, we developed and characterized a tool to interfere with Sema3A signaling by means of AAV-mediated expression of receptor bodies, soluble proteins formed by the extracellular domain of the endogenous Sema3A receptor (neuropilin1) fused to a human IgG Fc fragment. By using this tool to antagonize Sema3A signaling in the adult rat visual cortex, we found that the specific inhibition of Sema3A promoted ocular dominance plasticity. Thus, Sema3A accumulates in perineuronal nets in an experience-dependent manner and its presence in the mature visual cortex inhibits plasticity. [ABSTRACT FROM AUTHOR]

The most common neurodegenerative diseases are Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease, frontotemporal lobar degeneration, and the motor neuron diseases, with AD affecting approximately 6% of people aged 65 years and older, and PD affecting approximately 1% of people aged over 60 years. Specific proteins are associated with these neurodegenerative diseases, as determined by both immunohistochemical studies on post-mortem tissue and genetic screening, where protein misfolding and aggregation are key hallmarks. Many of these proteins are shown to misfold and aggregate into soluble non-native oligomers and large insoluble protein deposits (fibrils and plaques), both of which may exert a toxic gain of function. Proteotoxicity has been examined intensively in cell culture and in in vivo models, and clinical trials of methods to attenuate proteotoxicity are relatively new. Therapies to enhance cellular protein quality control mechanisms such as upregulation of chaperones and clearance/degradation pathways, as well as immunotherapies against toxic protein conformations, are being actively pursued. In this article, we summarize the common pathophysiology of neurodegenerative disease, and review therapies in early-phase clinical trials that target the proteotoxic component of several neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Determining protein thermal stability is integral in biomedical research. Here, with the use of two thermal stability assays, we show the melting temperature of amyloid precursor protein, an Alzheimer's disease related protein. The average melting temperature for amyloid precursor protein of 55.9 °C was derived from differential scanning fluorometry (55.1 ± 0.3 °C) and cellular thermal melt (56.7 ± 0.7 °C). These experimental methods have significant application for Alzheimer's disease research including their use for amyloid precursor protein stability profiling and for the identification of additional binding partners to further elucidate novel protein functions. [ABSTRACT FROM AUTHOR]

Puberty is driven by sophisticated neuroendocrine networks that timely activate the brain centers governing the reproductive axis. The timing of puberty is genetically determined; yet, puberty is also sensitive to numerous internal and external cues, among which metabolic/nutritional signals are especially prominent. Compelling epidemiological evidence suggests that alterations of the age of puberty are becoming more frequent; the underlying mechanisms remain largely unknown, but the escalating prevalence of obesity and other metabolic/feeding disorders is possibly a major contributing factor. This phenomenon may have clinical implications, since alterations in pubertal timing have been associated to adverse health outcomes, including higher risk of earlier all-cause mortality. This urges for a better understanding of the neurohormonal basis of normal puberty and its deviations. Compelling evidence has recently documented the master role of hypothalamic neurons producing kisspeptins, encoded by Kiss1, in the neuroendocrine pathways controlling puberty. Kiss1 neurons seemingly participate in transmitting the regulatory actions of metabolic cues on pubertal maturation. Key cellular metabolic sensors, as the mammalian target of rapamycin (mTOR), AMP-activated protein kinase (AMPK) and the fuel-sensing deacetylase, SIRT1, have been recently shown to participate also in the metabolic modulation of puberty. Recently, we have documented that AMPK and SIRT1 operate as major molecular effectors for the metabolic control of Kiss1 neurons and, thereby, puberty onset. Alterations of these molecular pathways may contribute to the perturbation of pubertal timing linked to conditions of metabolic stress in humans, such as subnutrition or obesity and might become druggable targets for better management of pubertal disorders. [ABSTRACT FROM AUTHOR]

Machado--Joseph disease (MJD), also known as spinocerebellar ataxia type 3 (SCA3), is an incurable disorder, widely regarded as the most common form of spinocerebellar ataxia in the world. MJD/SCA3 arises from mutation of the ATXN3 gene, but this simple monogenic cause contrasts with the complexity of the pathogenic mechanisms that are currently admitted to underlie neuronal dysfunction and death. The aberrantly expanded protein product -- ataxin-3 -- is known to aggregate and generate toxic species that disrupt several cell systems, including autophagy, proteostasis, transcription, mitochondrial function and signalling. Over the years, research into putative therapeutic approaches has often been devoted to the development of strategies that counteract disease at different stages of cellular pathogenesis. Silencing the pathogenic protein, blocking aggregation, inhibiting toxic proteolytic processing and counteracting dysfunctions of the cellular systems affected have yielded promising ameliorating results in studies with cellular and animal models. The current review analyses the available studies dedicated to the investigation of MJD/SCA3 pathogenesis and the exploration of possible therapeutic strategies, focusing primarily on gene therapy and pharmacological approaches rooted on the molecular and cellular mechanisms of disease. [ABSTRACT FROM AUTHOR]

Alzheimer’s disease (AD) is the most common neurodegenerative disease. Pathological proteins of AD mainly contain amyloid-beta (Aβ) and tau. Their deposition will lead to neuron damage by a series of pathways, and then induce memory and cognitive impairment. Thus, it is pivotal to understand the clearance pathways of Aβ and tau in order to delay or even halt AD. Aβ clearance mechanisms include ubiquitin-proteasome system, autophagy-lysosome, proteases, microglial phagocytosis, and transport from the brain to the blood via the blood-brain barrier (BBB), arachnoid villi and blood-CSF barrier, which can be named blood circulatory clearance. Recently, lymphatic clearance has been demonstrated to play a key role in transport of Aβ into cervical lymph nodes. The discovery of meningeal lymphatic vessels is another direct evidence for lymphatic clearance in the brain. Furthermore, periphery clearance also contributes to Aβ clearance. Tau clearance is almost the same as Aβ clearance. In this review, we will mainly introduce the clearance mechanisms of Aβ and tau proteins, and summarize corresponding targeted drug therapies for AD. [ABSTRACT FROM AUTHOR]

Amyloid deposition and neurofibrillary degeneration in Alzheimer's disease specifically affect discrete neuronal systems, but the underlying mechanisms that render some brain regions more vulnerable to Alzheimer's disease pathology than others remain largely unknown. Here we studied molecular properties underlying these distinct regional vulnerabilities by analysing Alzheimer's disease-typical neuroimaging patterns of amyloid deposition and neurodegeneration in relation to regional gene expression profiles of the human brain. Graded patterns of brain-wide vulnerability to amyloid deposition and neurodegeneration in Alzheimer's disease were estimated by contrasting multimodal amyloid-sensitive PET and structural MRI data between patients with Alzheimer's disease dementia (n = 76) and healthy controls (n = 126) enrolled in the Alzheimer's Disease Neuroimaging Initiative (ADNI). Regional gene expression profiles were derived from brain-wide microarray measurements provided by the Allen brain atlas of the adult human brain transcriptome. In a hypothesis-driven analysis focusing on the genes coding for the amyloid precursor (APP) and tau proteins (MAPT), regional expression levels of APP were positively correlated with the severity of regional amyloid deposition (r = 0.44, P = 0.009), but not neurodegeneration (r = 0.01, P = 0.96), whereas the opposite pattern was observed for MAPT (neurodegeneration: r = 0.46, P = 0.006; amyloid: r = 0.08, P = 0.65). Using explorative gene set enrichment analysis, amyloid-vulnerable regions were found to be characterized by relatively low expression levels of gene sets implicated in protein synthesis and mitochondrial respiration. By contrast, neurodegeneration-vulnerable regions were characterized by relatively high expression levels of gene sets broadly implicated in neural plasticity, with biological functions ranging from neurite outgrowth and synaptic contact over intracellular signalling cascades to proteoglycan metabolism. At the individual gene level this data-driven analysis further corroborated the association between neurodegeneration and MAPT expression, and additionally identified associations with known tau kinases (CDK5, MAPK1/ERK2) alongside components of their intracellular (Ras-ERK) activation pathways. Sensitivity analyses showed that these pathology-specific imaging-genetic associations were largely robust against changes in some of the methodological parameters, including variation in the brain donor sample used for estimating regional gene expression profiles, and local variations in the Alzheimer's disease-typical imaging patterns when these were derived from an independent patient cohort (BioFINDER study). These findings highlight that the regionally selective vulnerability to Alzheimer's disease pathology relates to specific molecular-functional properties of the affected neural systems, and that the implicated biochemical pathways largely differ for amyloid accumulation versus neurodegeneration. The data provide novel insights into the complex pathophysiological mechanisms of Alzheimer's disease and point to pathology-specific treatment targets that warrant further exploration in independent studies. [ABSTRACT FROM AUTHOR]

One characteristic of Alzheimer’s disease (AD) is excessive accumulation of amyloid-β (Aβ) in the brain. Aβ brain accumulation is, in part, due to a reduction in Aβ clearance from the brain across the blood-brain barrier. One key element that contributes to Aβ brain clearance is P-glycoprotein (P-gp) that transports Aβ from brain to blood. In AD, P-gp protein expression and transport activity levels are significantly reduced, which impairs Aβ brain clearance. The mechanism responsible for reduced P-gp expression and activity levels is poorly understood. We recently demonstrated that Aβ40 triggers P-gp degradation through the ubiquitin-proteasome pathway. Consistent with these data, we show here that ubiquitinated P-gp levels in brain capillaries isolated from brain samples of AD patients are increased compared to capillaries isolated from brain tissue of cognitive normal individuals. We extended this line of research to in vivo studies using transgenic human amyloid precursor protein (hAPP)-overexpressing mice (Tg2576) that were treated with PYR41, a cell-permeable, irreversible inhibitor of the ubiquitin-activating enzyme E1. Our data show that inhibiting P-gp ubiquitination protects the transporter from degradation, and immunoprecipitation experiments confirmed that PYR41 prevented P-gp ubiquitination. We further found that PYR41 treatment prevented reduction of P-gp protein expression and transport activity levels and substantially lowered Aβ brain levels in hAPP mice. Together, our findings provide in vivo proof that the ubiquitin-proteasome system mediates reduction of blood-brain barrier P-gp in AD and that inhibiting P-gp ubiquitination prevents P-gp degradation and lowers Aβ brain levels. Thus, targeting the ubiquitin-proteasome system may provide a novel therapeutic approach to protect blood-brain barrier P-gp from degradation in AD and other Aβ-based pathologies. [ABSTRACT FROM AUTHOR]

The ubiquitin-proteasome system (UPS) is the main pathway responsible for the degradation of misfolded proteins, and its dysregulation has been implicated in several neurodegenerative diseases, including Alzheimer's disease (AD). UBB+1, a mutant variant of ubiquitin B, was found to accumulate in neurons of AD patients and it has been linked to UPS dysfunction and neuronal death. Using the yeast Saccharomyces cerevisiae as a model system, we constitutively expressed UBB+1 to evaluate its effects on proteasome function and cell death, particularly under conditions of chronological aging. We showed that the expression of UBB+1 caused inhibition of the three proteasomal proteolytic activities (caspase-like (b1), trypsin-like (b2) and chymotrypsin-like (b5) activities) in yeast. Interestingly, this inhibition did not alter cell viability of growing cells. Moreover, we showed that cells expressing UBB+1 at lower level displayed an increased capacity to degrade induced misfolded proteins. When we evaluated cells during chronological aging, UBB+1 expression at lower level, prevented cells to accumulate reactive oxygen species (ROS) and avert apoptosis, dramatically increasing yeast life span. Since proteasome inhibition by UBB+1 has previously been shown to induce chaperone expression and thus protect against stress, we evaluated our UBB+1 model under heat shock and oxidative stress. Higher expression of UBB+1 caused thermotolerance in yeast due to induction of chaperones, which occurred to a lesser extent at lower expression level of UBB+1 (where we observed the phenotype of extended life span). Altering UPS capacity by differential expression of UBB+1 protects cells against several stresses during chronological aging. This system can be valuable to study the effects of UBB+1 on misfolded proteins involved in neurodegeneration and aging. [ABSTRACT FROM AUTHOR]

Alzheimer disease (AD) pathology includes the accumulation of poly-ubiquitylated (also known as poly-ubiquitinated) proteins and failures in proteasome-dependent degradation. Whereas the distribution of proteasomes and its role in synaptic function have been studied, whether proteasome activity regulates the axonal transport and metabolism of the amyloid precursor protein (APP), remains elusive. By using live imaging in primary hippocampal neurons, we showed that proteasome inhibition rapidly and severely impairs the axonal transport of APP. Fluorescence cross-correlation analyses andmembrane internalization blockage experiments showed that plasma membrane APP does not contribute to transport defects. Moreover, by western blotting and double-color APP imaging, we demonstrated that proteasome inhibition precludes APP axonal transport by enhancing its endo-lysosomal delivery, where ß-cleavage is induced. Taken together, we found that proteasomes control the distal transport of APP and can re-distribute Golgi-derived vesicles to the endo-lysosomal pathway. This crosstalk between proteasomes and lysosomes regulates the intracellular APP dynamics, and defects in proteasome activity can be considered a contributing factor that leads to abnormal APP metabolism in AD. [ABSTRACT FROM AUTHOR]

Polyglutamine (polyQ) diseases are hereditary neurodegenerative disorders caused by an abnormal expansion of a trinucleotide CAG repeat in the coding region of their respective associated genes. PolyQ diseases mainly display progressive degeneration of the brain and spinal cord. Nine polyQ diseases are known, including Huntington's disease (HD), spinal and bulbar muscular atrophy (SBMA), dentatorubralpallidoluysian atrophy (DRPLA), and six forms of spinocerebellar ataxia (SCA). HD is the best characterized polyQ disease. Many studies have reported that transcriptional dysregulation and post-translational disruptions, which may interact with each other, are central features of polyQ diseases. Post-translational modifications, such as the acetylation of histones, are closely associated with the regulation of the transcriptional activity. A number of groups have studied the interactions between the polyQ proteins and transcription factors. Pharmacological drugs or genetic manipulations aimed at correcting the dysregulation have been confirmed to be effective in the treatment of polyQ diseases in many animal and cellular models. For example, histone deaceylase inhibitors have been demonstrated to have beneficial effects in cases of HD, SBMA, DRPLA, and SCA3. In this review, we describe the transcriptional and post-translational dysregulation in polyQ diseases with special focus on HD, and we summarize and comment on potential treatment approaches targeting disruption of transcription and post-translation processes in these diseases. [ABSTRACT FROM AUTHOR]

Guam parkinsonism-dementia complex (G-PDC) is an enigmatic neurodegenerative disease that is endemic to the Pacific island of Guam. G-PDC patients are clinically characterized by progressive cognitive impairment and parkinsonism. Neuropathologically, G-PDC is characterized by abundant neurofibrillary tangles, which are composed of hyperphosphorylated tau, marked deposition of 43-kDa TAR DNA-binding protein, and neuronal loss. Although both genetic and environmental factors have been implicated, the etiology and pathogenesis of G-PDC remain unknown. Recent neuropathological studies have provided new clues about the pathomechanisms involved in G-PDC. For example, deposition of abnormal components of the protein quality control system in brains of G-PDC patients indicates a role for proteostasis imbalance in the disease. This opens up promising avenues for new research on G-PDC and could have important implications for the study of other neurodegenerative disorders. [ABSTRACT FROM AUTHOR]

ABSTRACT The relationship between atmospheric circulation and climate in Svalbard has been described in dozens of studies. However, the data used for that purpose usually came from permanent stations on the coast. The influence of atmospheric circulation on topoclimatic diversity has not been explored so often, and hardly at all for other periods than the Arctic summer. In this article, the relationships between circulation and air temperature are described using daily data sourced from six sites located around Forlandsundet ( NW Spitsbergen) during 2010-2013. The analysis was conducted independently for three seasons identified as: winter (Nov-Mar), spring/autumn (Apr-May and Sep-Oct) and summer (Jun-Aug) and also for three air temperature parameters: diurnal mean (Ti), maximum (Tmax) and minimum (Tmin) temperature. The atmospheric circulation in the studied area was described using Tadeusz Niedźwiedź's classification of diurnal circulation types for Svalbard. The influence of atmospheric circulation on the spatial pattern of air temperature is not uniform across the Forlandsundet region; in particular, important differences were observed between coastal and inland parts of the study area. Thus, generalization of relationships between air temperature and atmospheric circulation for the entire area of Spitsbergen based on data only from coastal stations is not appropriate. The influence of atmospheric circulation on the spatial pattern of air temperature in the Forlandsundet region also changes through the year. In the cold season (Sep-May) it differs significantly from that observed in summer (Jun-Aug), and this feature is also seen in analyses of the 10% highest (≥ 90th percentile) and lowest (≤ 10th percentile) thermal differences. In summer, the influence of atmospheric circulation on air temperature in the topoclimatic scale is definitely less stable than in the cold season. [ABSTRACT FROM AUTHOR]

Ubiquitination of proliferating cell nuclear antigen (PCNA) plays an important role in DNA damage response. Ectopic expression of PCNA fused at either terminus with ubiquitin (Ub) lacking two C-terminal glycine residues induces translesion DNA synthesis which resembles synthesis mediated by PCNA monoubiquitination. PCNA fused with Ub containing the C-terminal Gly residues at the C-terminus can be further polyubiquitinated in a Gly-dependent manner, which inhibits cell proliferation and induces ATR-dependent replication checkpoint. In this study, we surprisingly found that PCNA fused to a head-to-tail linear Ub chain induces apoptosis in a Ub chain length-dependent manner. Further investigation revealed that the apoptotic effect is actually induced by the linear Ub chain independently from PCNA, as the Ub chain fused to GFP or an epitope tag still efficiently induces apoptosis. It is revealed that the artificial linear Ub chain differs from endogenously encoded linear Ub chains in that its Ubs contain a Ub-G76S substitution, making the Ub chain resistant to cleavage by deubiquitination enzymes. We demonstrated in this study that ectopic expression of the artificial Ub chain alone in cultured human cancer cells is sufficient to inhibit tumor growth in a xenograft mouse model, making the linear Ub chain a putative anti-cancer agent. [ABSTRACT FROM AUTHOR]

Congenital hypogonadotropic hypogonadism ( CHH) is a rare genetic form of isolated gonadotropin-releasing hormone (Gn RH) deficiency caused by mutations in > 30 genes. Fibroblast growth factor receptor 1 ( FGFR1) is the most frequently mutated gene in CHH and is implicated in Gn RH neuron development and maintenance. We note that a CHH FGFR1 mutation (p.L342S) decreases signaling of the metabolic regulator FGF21 by impairing the association of FGFR1 with β-Klotho ( KLB), the obligate co-receptor for FGF21. We thus hypothesized that the metabolic FGF21/ KLB/ FGFR1 pathway is involved in CHH. Genetic screening of 334 CHH patients identified seven heterozygous loss-of-function KLB mutations in 13 patients (4%). Most patients with KLB mutations (9/13) exhibited metabolic defects. In mice, lack of Klb led to delayed puberty, altered estrous cyclicity, and subfertility due to a hypothalamic defect associated with inability of Gn RH neurons to release Gn RH in response to FGF21. Peripheral FGF21 administration could indeed reach Gn RH neurons through circumventricular organs in the hypothalamus. We conclude that FGF21/ KLB/ FGFR1 signaling plays an essential role in Gn RH biology, potentially linking metabolism with reproduction. [ABSTRACT FROM AUTHOR]

Tumor progression alters the composition and physical properties of the extracellular matrix. Particularly, increased matrix stiffness has profound effects on tumor growth and metastasis. While endothelial cells are key players in cancer progression, the influence of tumor stiffness on the endothelium and the impact on metastasis is unknown. Through quantitative mass spectrometry, we find that the matricellular protein CCN1/ CYR61 is highly regulated by stiffness in endothelial cells. We show that stiffness-induced CCN1 activates β-catenin nuclear translocation and signaling and that this contributes to upregulate N-cadherin levels on the surface of the endothelium, in vitro. This facilitates N-cadherin-dependent cancer cell-endothelium interaction. Using intravital imaging, we show that knockout of Ccn1 in endothelial cells inhibits melanoma cancer cell binding to the blood vessels, a critical step in cancer cell transit through the vasculature to metastasize. Targeting stiffness-induced changes in the vasculature, such as CCN1, is therefore a potential yet unappreciated mechanism to impair metastasis. [ABSTRACT FROM AUTHOR]

The ubiquitin-proteasome system (UPS) is one of the major mechanisms for protein breakdown in cells, targeting proteins for degradation by enzymatically conjugating them to ubiquitin molecules. Intracellular accumulation of ubiquitin-B (UBB), a frameshift mutant of ubiquitin-B, is indicative of a dysfunctional UPS and has been implicated in several disorders, including neurodegenerative disease. UBB-expressing transgenic mice display widespread labeling for UBB in brain and exhibit behavioral deficits. Here, we show that UBB is specifically expressed in a subset of parasagittal stripes of Purkinje cells in the cerebellar cortex of a UBB-expressing mouse model. This expression pattern is reminiscent of that of the constitutively expressed Purkinje cell antigen HSP25, a small heat shock protein with neuroprotective properties. [ABSTRACT FROM AUTHOR]

Terminal Schwann cells (TSCs) are specialized cells that envelop the motor nerve terminal, and play a role in the maintenance and regeneration of neuromuscular junctions (NMJs). The chemorepulsive protein semaphorin 3A (SEMA3A) is selectively up-regulated in TSCs on fast-fatigable muscle fibers following experimental denervation of the muscle (BotoxA-induced paralysis or crush injury to the sciatic nerve) or in the motor neuron disease amyotrophic lateral sclerosis (ALS). Re-expression of SEMA3A in this subset of TSCs is thought to play a role in the selective plasticity of nerve terminals as observed in ALS and following BotoxA-induced paralysis. Using a mouse model expressing a mutant SEMA3A with diminished signaling capacity, we studied the influence of SEMA3A signaling at the NMJ with two denervation paradigms; a motor neuron disease model (the G93A-hSOD1 ALS mouse line) and an injury model (BotoxA-induced paralysis). ALS mice that either expressed 1 or 2 mutant SEMA3A alleles demonstrated no difference in ALS-induced decline in motor behavior. We also investigated the effects of BotoxA-induced paralysis on the sprouting capacity of NMJs in the K108N-SEMA3A mutant mouse, and observed no change in the differential neuronal plasticity found at NMJs on fast-fatigable or slow muscle fibers due to the presence of the SEMA3A mutant protein. Our data may be explained by the residual repulsive activity of the mutant SEMA3A, or it may imply that SEMA3A alone is not a key component of the molecular signature affecting NMJ plasticity in ALS or BotoxA-induced paralysis. Interestingly, we did observe a sex difference in motor neuron sprouting behavior after BotoxA-induced paralysis in WT mice which we speculate may be an important factor in the sex dimorphic differences seen in ALS. [ABSTRACT FROM AUTHOR]

An important mechanism of neuronal plasticity is neurogenesis, which occurs during the embryonic period, forming the brain and its structure, and in the postnatal period, providing repair processes and participating in the mechanisms of memory consolidation. Adult neurogenesis in mammals, including humans, is limited in two specific brain areas, the lateral walls of the lateral ventricles (subventricular zone) and the granular layer of the dentate gyrus of the hippocampus (subgranular zone). Neural stem cells (NSC), self-renewing, multipotent progenitor cells, are formed in these zones. Neural stem cells are capable of differentiating into the basic cell types of the nervous system. In addition, NSC may have neurogenic features and non-specific non-neurogenic functions aimed at maintaining the homeostasis of the brain. The microenvironment formed in neurogenic niches has importance maintaining populations of NSC and regulating differentiation into neural or glial cells via cell-to-cell interactions and microenvironmental signals. The vascular microenvironment in neurogenic niches are integrated by signaling molecules secreted from endothelial cells in the blood vessels of the brain or by direct contact with these cells. Accumulation of astrocytes in neurogenic niches if also of importance and leads to activation of neurogenesis. Dysregulation of neurogenesis contributes to the formation of neurological deficits observed in neurodegenerative diseases. Targeting regulation of neurogenesis could be the basis of new protocols of neuroregeneration. [ABSTRACT FROM AUTHOR]

The ubiquitin mutant UBB+1 has been identified as a hallmark of neurodegenerative diseases. In this study, we characterize polyubiquitinated forms of UBB+1 in vitro and from patient samples. The ability of UBB+1 to be readily ubiquitinated by several E2 enzymes provided a mechanism for the controlled synthesis and purification of defined conjugates. This allowed us to utilize polyUb- UBB+1 conjugates for biochemical assays, as well as solution NMR. Coupled with our immunoassay for detection of ubiquitinated forms of UBB+1 in patient blood samples, we gain a clearer picture of the molecular mechanisms underlying neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Several integrated proteolytic systems contribute to the maintenance of cellular homeostasis through the continuous removal of misfolded, aggregated or oxidized proteins and damaged organelles. Among these systems, the proteasome and autophagy play the major role in protein quality control, which is a fundamental issue in non-proliferative cells such as neurons. Disturbances in the functionality of these two pathways are frequently observed in neurodegenerative diseases, like Alzheimer's disease, and reflect the accumulation of protease-resistant, deleterious protein aggregates. In this review, we explored the sophisticated crosstalk between the ubiquitin-proteasome system and autophagy in the removal of the harmful structures that characterize Alzheimer's disease neurons. We also dissected the role of the numerous shuttle factors and chaperones that, directly or indirectly interacting with ubiquitin and LC3, are used for cargo selection and delivery to one pathway or the other. [ABSTRACT FROM AUTHOR]

The median eminence (ME) of the hypothalamus comprises the hypothalamic nerve terminals, glia (especially tanycytes) and the portal capillary vasculature that transports hypothalamic neurohormones to the anterior pituitary gland. The ultrastructure of the ME is dynamically regulated by hormones and undergoes organizational changes during development and reproductive cycles in adult females, but relatively little is known about the ME during aging, especially in nonhuman primates. Therefore, we used a novel transmission scanning electron microscopy technique to examine the cytoarchitecture of the ME of young and aged female rhesus macaques in a preclinical monkey model of menopausal hormone treatments. Rhesus macaques were ovariectomized and treated for 2 years with vehicle, estradiol (E2), or estradiol + progesterone (E2 + P4). While the overall cytoarchitecture of the ME underwent relatively few changes with age and hormones, changes to some features of neural and glial components near the portal capillaries were observed.Specifically, large neuroterminal size was greater in aged compared to young adult animals, an effect that was mitigated or reversed by E2 alone but not by E2 + P4 treatment. Overall glial size and the density and tissue fraction of the largest subset of glia were greater in aged monkeys, and in some cases reversed by E2 treatment. Mitochondrial size was decreased by E2 , but not E2 + P4 , only in aged macaques. These results contrast substantially with work in rodents, suggesting that the ME of aging macaques is less vulnerable to age-related disorganization, and that the effects of E2 on monkeys' ME are age specific. [ABSTRACT FROM AUTHOR]

The proteasome is a structural complex of many proteins that degrades substrates marked by covalent linkage to ubiquitin. Many years of research has shown a role for ubiquitin-proteasome-mediated proteolysis in synaptic plasticity and memory mainly in degrading synaptic, cytoplasmic and nuclear proteins. Recent work indicates that the proteasome has wider proteolytic and non-proteolytic roles in processes such as histone modifications that affect synaptic plasticity and memory. In this review, we assess the evidence gathered from neuronal as well as non-neuronal cell types regarding the function of the proteasome in positive or negative regulation of posttranslational modifications of histones, such as acetylation, methylation and ubiquitination. We discuss the critical roles of the proteasome in clearing excess histone proteins in various cellular contexts and the possible nonproteolytic functions in regulating transcription of target genes. In addition, we summarize the current literature on diverse chromatin-remodeling machineries, such as histone acetyltransferases, deacetylates, methyltransferases and demethylases, as targets for proteasomal degradation across experimental models. Lastly, we provide a perspective on how proteasomal regulation of histone modifications may modulate synaptic plasticity in the nervous system. [ABSTRACT FROM AUTHOR]

Alzheimer's disease ( AD) is the most common neurodegenerative disease and is characterized by accumulation of misfolded and aggregated proteins. Since the ubiquitin-proteasome system ( UPS) is the major intracellular protein quality control ( PQC) system in eukaryotic cells, it is likely involved in the etiology of AD. The frameshift form of ubiquitin (Ubb+1) accumulates in the neuritic plaques and neurofibrillary tangles in patients with AD. Ubb+1 accumulates in an age-dependent manner as a result of RNA-polymerase mediated molecular misreading during transcription, which allows the formation of mutant proteins in the absence of gene mutations. The accumulation of the Ubb+1 protein may act as an endogenous reporter for proteasome dysfunction and a growing number of studies have shown that Ubb+1 may play more important pathogenic roles in AD etiology than previously hypothesized. The yeast Saccharomyces cerevisiae shares many conserved biological processes with all eukaryotic cells, including human neurons. This organism has been regarded as a model system for investigating the fundamental intracellular mechanisms, including those underlying neurodegeneration. We propose here that yeast systems biology approaches, combined with cell and molecular biology approaches will increase the relevant knowledge needed for advancement and elucidation of mechanisms and complex traits, which could provide new targets for therapeutic intervention in AD. WIREs Syst Biol Med 2016, 8:300-313. doi: 10.1002/wsbm.1340 For further resources related to this article, please visit the . [ABSTRACT FROM AUTHOR]

The spontaneous mutation tambaleante is caused by the Gly483Glu substitution in the highly conserved N terminal RCC1-like domain of the HERC1 protein, which leads to the increase of mutated protein levels responsible for cerebellar Purkinje cell death by autophagy. Until now, Purkinje cells have been the only central nervous neurons reported as being targeted by the mutation, and their degeneration elicits an ataxic syndrome in adult mutant mice. However, the ultrastructural analysis performed here demonstrates that signs of autophagy, such as autophagosomes, lysosomes, and altered mitochondria, are present in neocortical pyramidal, CA3 hippocampal pyramidal, and spinal cord motor neurons. The main difference is that the reduction in the number of neurons affected in the tambaleante mutation in the neocortex, the hippocampus, and the spinal cord is not so evident as the dramatic loss of cerebellar Purkinje cells. Interestingly, signs of autophagy are absent in both interneurons and neuroglia cells. Affected neurons have in common that they are projection neurons which receive strong and varied synaptic inputs, and possess the highest degree of neuronal activity. Therefore, because the integrity of the ubiquitin-proteasome system is essential for protein degradation and hence, for normal protein turnover, it could be hypothesized that the deleterious effects of the misrouting of these pathways would depend directly on the neuronal activity. [ABSTRACT FROM AUTHOR]

Aims Polyglutamine ( polyQ) diseases are characterized by the expansion of a polymorphic glutamine sequence in disease-specific proteins and exhibit aggregation of these proteins. This is combated by the cellular protein quality control ( PQC) system, consisting of chaperone-mediated refolding as well as proteasomal and lysosomal degradation pathways. Our recent study in the polyQ disease spinocerebellar ataxia type 3 ( SCA3) suggested a distinct pattern of protein aggregation and PQC dysregulation. Methods To corroborate these findings we have investigated immunohistochemically stained 5 μm sections from different brain areas of Huntington's disease ( HD) and SCA3 patients. Results Irrespective of disease and brain region, we observed peri- and intranuclear polyQ protein aggregates. A subset of neurones with intranuclear inclusions bodies exhibited signs of proteasomal dysfunction, up-regulation of HSPA1A and re-distribution of DNAJB1. The extent of the observed effects varied depending on brain area and disease protein. Conclusions Our results suggest a common sequence, in which formation of cytoplasmic and nuclear inclusions precede proteasomal impairment and induction of the cellular stress response. Clearly, impairment of the PQC is not the primary cause for inclusion formation, but rather a consequence that might contribute to neuronal dysfunction and death. Notably, the inclusion pathology is not directly correlated to the severity of the degeneration in different areas, implying that different populations of neurones respond to polyQ aggregation with varying efficacy and that protein aggregation outside the neuronal perikaryon (e.g. axonal aggregates) or other effects of polyQ aggregation, which are more difficult to visualize, may contribute to neurodegeneration. [ABSTRACT FROM AUTHOR]

Macroautophagy and chaperone-mediated autophagy (CMA) are two important subtypes of autophagy that play a critical role in cellular quality control under physiological and pathological conditions. Despite the marked differences between these two autophagic pathways, macroautophagy and CMA are intimately connected with each other during the autophagy-lysosomal degradation process, in particular, in the setting of neurological illness. Macroautophagy serves as a backup mechanism to removal of malfunctioning proteins (i.e., aberrant α-synuclein) from the cytoplasm when CMA is compromised, and vice versa. The molecular mechanisms underlying the conversation between macroautophagy and CMA are being clarified. Herein, we survey current overviews concentrating on the complex interactions between macroautophagy and CMA, and present therapeutic potentials through utilization and manipulation of macroautophagy-CMA crosstalk in the treatment of neurological diseases. [ABSTRACT FROM AUTHOR]

The article discusses the availability of pharmacological interventions for the treatment of Parkinson's disease (PD). It states that pharmacological dopamine replacement considered to be main therapy for treating PD which is caused mainly by progressive loss of dopaminergic neurons in the substantia nigra par compacta (SNpc) owing to one of the axon guidance molecule known as repulsive guidance molecule a (RGMa). It mentions therapies are formulated to tackle all possibilities including RGMa.

The semaphorin proteins, which contribute to the morphogenesis and homeostasis of a wide range of systems, are among the best-studied families of guidance cues. Much recent research has focused on the role of semaphorins in the development and adult activity of hormone systems and, reciprocally, how circulating reproductive hormones regulate their expression and function. Specifically, several reports have focused on the molecular mechanisms underlying the effects of semaphorins on the migration, survival and structural and functional plasticity of neurons that secrete gonadotropin-releasing hormone (GnRH), essential for the acquisition and maintenance of reproductive competence in mammals. Alterations in the development of this neuroendocrine system lead to anomalous or absent GnRH secretion, resulting in heterogeneous reproductive disorders such as congenital hypogonadotropic hypogonadism (CHH) or other conditions characterized by infertility or subfertility. This review summarizes current knowledge of the role of semaphorins and their receptors on the development, differentiation and plasticity of the GnRH system. In addition, the involvement of genetic deficits in semaphorin signaling in some forms of CHH in humans is discussed. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Amyloid β (Aβ) plaque formation is a prominent cellular hallmark of Alzheimer's disease (AD). To date, immunization trials in AD patients have not been effective in terms of curing or ameliorating dementia. In addition, γ-secretase inhibitor strategies await clinical improvements in AD. These approaches were based upon the idea that autosomal dominant mutations in amyloid precursor protein (APP) and Presenilin 1 (PS1) genes are predictive for treatment of all AD patients. However most AD patients are of the sporadic form which partly explains the failures to treat this multifactorial disease. The major risk factor for developing sporadic AD (SAD) is aging whereas the Apolipoprotein E polymorphism (ε4 variant) is the most prominent genetic risk factor. Other medium-risk factors such as triggering receptor expressed on myeloid cells 2 (TREM2) and nine low risk factors from Genome Wide Association Studies (GWAS) were associated with AD. Recently, pooled GWAS studies identified protein ubiquitination as one of the key modulators of AD. In addition, a brain site specific strategy was used to compare the proteomes of AD patients by an Ingenuity Pathway Analysis. This strategy revealed numerous proteins that strongly interact with ubiquitin (UBB) signaling, and pointing to a dysfunctional ubiquitin proteasome system (UPS) as a causal factor in AD. We reported that DNA-RNA sequence differences in several genes including ubiquitin do occur in AD, the resulting misframed protein of which accumulates in the neurofibrillary tangles (NFTs). This suggests again a functional link between neurodegeneration of the AD type and loss of protein quality control by the UPS. Progress in this field is discussed and modulating the activity of the UPS opens an attractive avenue of research towards slowing down the development of AD and ameliorating its effects by discovering prime targets for AD therapeutics. [ABSTRACT FROM AUTHOR]

The ubiquitin-proteasome system (UPS) plays a fundamental role in protein degradation in neurons, and there is strong evidence that it fulfills a key role in synaptic transmission. The aim of the present work was to study the implication of one component of the UPS, the HERC1 E3 Ubiquitin Ligase, in motor function and neuromuscular transmission. The tambaleante ( tbl) mutant mouse carries a spontaneous mutation in HERC1 E3 Ubiquitin Ligase, provoking an ataxic phenotype that develops in the second month of life. Our results show that motor performance in mutant mice is altered at postnatal day 30, before the cerebellar neurodegeneration takes place. This defect is associated with by: (a) a reduction of the motor end-plate area, (b) less efficient neuromuscular activity in vivo, and (c) an impaired evoked neurotransmitter release. Together, these data suggest that the HERC1 E3 Ubiquitin Ligase is fundamental for normal muscle function and that it is essential for neurotransmitter release at the mouse neuromuscular junction. [ABSTRACT FROM AUTHOR]

Background: Osteoblastogenesis is under delicate control by multiple factors and hormones. Recent reports indicated the involvement of immunological and neuronal regulators. However, the role of neuropilin 1 (Nrp1) in osteoblastogenesis remains obscure. Methods: Real-time PCR was carried out to detect the mRNA of osteoblastic markers, Nrp1, and discoidin domain receptor 2 (DDR2). Immunoblot was performed to test the protein of Nrp1 and DDR2. Osteogenic differentiation was evaluated by mRNA analysis of osteogenic markers, and determination of ALP activity and OCN secretion. The intercellular signaling effectors were examined by immunoblot. Immunofluorescent assays were performed to detect the localization of Nrp1 and DDR2. Half-life determination assay was executed to test the DDR2 stability. Results: The expression of Nrp1 paralleled with that of DDR2 during osteoblastogensis. Nrp1 overexpression enhanced DDR2-induced stimulation of osteoblastogensis, whereas Nrp1 silencing caused attenuation. Nrp1 overexpression increased the phosphorylation of DDR2, ERK1/2 and Runx2. Nrp1 co-localized with DDR2 in the cellular membrane of differentiated MC3T3-E1. Enhanced or attenuated Nrp1 expression did not alter the mRNA transcript of DDR2. Nrp1 overexpression prolonged the half-life of DDR2 protein. Conclusion: Our results originally demonstrated the stimulatory role of Nrp1 in DDR2-induced osteoblast differentiation, providing molecular evidence for exploiting Nrp1 and DDR2 as targets to treat bone-related disease. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Critically impaired protein degradation is discussed to contribute to neurodegenerative disorders, including Parkinson's, Huntington's, Alzheimer's, and motor neuron diseases. Misfolded, aggregated, or surplus proteins are efficiently degraded via distinct protein degradation pathways, including the ubiquitin-proteasome system, autophagy, and vesicular trafficking. These pathways are regulated by covalent modification of target proteins with the small protein ubiquitin and are evolutionary highly conserved from humans to yeast. The yeast Saccharomyces cerevisiae is an established model for deciphering mechanisms of protein degradation, and for the elucidation of pathways underlying programmed cell death. The expression of human neurotoxic proteins triggers cell death in yeast, with neurotoxic protein-specific differences. Therefore, yeast cell death models are suitable for analyzing the role of protein degradation pathways in modulating cell death upon expression of disease-causing proteins. This review summarizes which protein degradation pathways are affected in these yeast models, and how they are involved in the execution of cell death. I will discuss to which extent this mimics the situation in other neurotoxic models, and how this may contribute to a better understanding of human disorders. [ABSTRACT FROM AUTHOR]

Loss of protein quality control by the ubiquitin-proteasome system (UPS) during aging is one of the processes putatively contributing to cellular stress and Alzheimer's disease (AD) pathogenesis. Recently, pooled GenomeWide Association Studies (GWAS), pathway analysis and proteomics identified protein ubiquitination as one of the key modulators of AD. Mutations in ubiquitin B mRNA that result in UBB+1 dose-dependently cause an impaired UPS, subsequent accumulation of UBB+1 and most probably depositions of other aberrant proteins present in plaques and neurofibrillary tangles. We used specific immunohistochemical probes for a comprehensive topographic mapping of the UBB+1 distribution in the brains of transgenic mouse line 3413 overexpressing UBB+1.We also mapped the expression of UBB+1 in brain areas of AD patients selected based upon the distribution of UBB+1 in line 3413. Therefore, we focused on the olfactory bulb, basal ganglia, nucleus basalis of Meynert, inferior colliculus and raphe nuclei. UBB+1 distribution was compared with established probes for pre-tangles and tangles and Ab plaques. UBB+1 distribution found in line 3413 is partly mirrored in the AD brain. Specifically, nuclei with substantial accumulations of tangle-bearing neurons, such as the nucleus basalis of Meynert and raphe nuclei also present high densities of UBB+1 positive tangles. Line 3413 is useful for studying the contribution of proteasomal dysfunction in AD. The findings are consistent with evidence that areas outside the forebrain are also affected in AD. Line 3413 may also be predictive for other conformational diseases, including related tauopathies and polyglutamine diseases, in which UBB+1 accumulates in their cellular hallmarks. [ABSTRACT FROM AUTHOR]

Mitral and tufted cells are the projection neurons of the olfactory bulb ( OB). We previously reported that somata location and innervation patterns were different between early- and late-born mitral cells (Imamura et al., 2011). Here, we introduced a plasmid that drives the expression of a GFP gene into the mouse OB using in utero electroporation, and demonstrated that we can deliver the plasmid vectors into distinct subsets of OB projection neurons by changing the timing of electroporation after fertilisation. The electroporation performed at embryonic day (E)10 preferentially labeled mitral cells in the accessory OB and main OB mitral cells in dorsomedial mitral cell layer ( MCL). In contrast, the E12 electroporation introduced the plasmid vectors preferentially into main OB mitral cells in the ventrolateral MCL and tufted cells. Combining these data with BrdU injections, we confirmed that E10 and E12 electroporation preferentially labeled early- and late-born projection neurons, respectively. This work introduces a novel method for segregated labeling of mouse olfactory bulb projection neurons based on their birthdates. With this technique we found that early- and late-born projection neurons extend their secondary dendrites in the deep and superficial external plexiform layer ( EPL), respectively. Although a similar segregation has been suggested for mitral vs. tufted cell dendrites, we found mitral cells projecting secondary dendrites into the superficial EPL in E12-electroporated main OB. Our observations indicate that timing of neurogenesis regulates not only somata location and innervation patterns but also the laminar organisation of projection neuron dendrites in the EPL. [ABSTRACT FROM AUTHOR]

The amyloid-β protein precursor/presenilin 1 (AβPP/PS1) mouse model of Alzheimer's disease (AD) has provided robust neuropathological hallmarks of familial AD-like pattern. AD is a neurodegenerative process that causes severe cognitive impairment; it is characterized by the accumulation of amyloid-β (Aβ) and hyperphosphorylated tau forms and by oxidative and inflammatory processes in brain. Currently, efforts are made to understand biochemical pathways because there is no effective therapy for AD. Resveratrol is a polyphenol that induces expression and activation of several neuroprotective pathways involving Sirtuin1 and AMPK. The objective of this work was to assess the effect of oral resveratrol administration on AβPP/PS1 mice. Long-term resveratrol treatment significantly prevented memory loss as measured by the object recognition test. Moreover, resveratrol reduced the amyloid burden and increased mitochondrial complex IV protein levels in mouse brain. These protective effects of resveratrol were mainly mediated by increased activation of Sirtuin 1 and AMPK pathways in mice. However, an increase has been observed in IL1β and TNF gene expression, indicating that resveratrol promoted changes in inflammatory processes, although no changes were detected in other key actors of the oxidative stress pathway. Taken together, our findings suggest that resveratrol is able to reduce the harmful process that occurs in AβPP/PS1 mouse hippocampus, preventing memory loss. [ABSTRACT FROM AUTHOR]

Spinocerebellar ataxia type 2 ( SCA2) is a hereditary neurodegenerative disorder caused by the expansion of the trinucleotide CAG repeats encoding elongated polyglutamine tract in ataxin-2, the SCA2 gene product. Polyglutamine diseases comprise nine genetic entities, including seven different forms of spinocerebellar ataxias, Huntington's disease, and spinal and bulbar muscular atrophy. These are pathologically characterized by neuronal loss and intranuclear aggregates or inclusions of mutant proteins including expanded polyglutamine in selected neuronal groups. Previously, we examined immunolocalization of ubiquitin, expanded polyglutamine (probed by 1 C2 antibody), and ataxin-2 in genetically confirmed SCA2 patients. In the present study, we expanded this approach by distinguishing different patterns of subcellular 1 C2 immunoreactivity ('granular cytoplasmic,' 'cytoplasmic and nuclear' and 'nuclear with inclusions.') and by quantifying their regional frequencies in three autopsied SCA2 brains at different stage of the disease. Comparison with neuronal loss and gliosis revealed that overall 1 C2 immunoreactivity was paralleled with their severity. Furthermore, appearance of granular cytoplasmic pattern corresponded to early stage, cytoplasmic and nuclear pattern to active stage, and nuclear with inclusions pattern to final stage. We conclude that this 1 C2-immunoreactive typing may be useful for evaluating the overall severity and extent of affected regions and estimating the neuropathological stage of SCA2. [ABSTRACT FROM AUTHOR]

Amyotrophic Lateral Sclerosis (ALS) is being redefined as a distal axonopathy, in that many molecular changes influencing motor neuron degeneration occur at the neuromuscular junction (NMJ) at very early stages of the disease prior to symptom onset. A huge variety of genetic and environmental causes have been associated with ALS, and interestingly, although the cause of the disease can differ, both sporadic and familial forms of ALS show a remarkable similarity in terms of disease progression and clinical manifestation. The NMJ is a highly specialized synapse, allowing for controlled signaling between muscle and nerve necessary for skeletal muscle function. In this review we will evaluate the clinical, animal experimental and cellular/molecular evidence that supports the idea of ALS as a distal axonopathy. We will discuss the early molecular mechanisms that occur at the NMJ, which alter the functional abilities of the NMJ. Specifically, we focus on the role of axon guidance molecules on the stability of the cytoskeleton and how these molecules may directly influence the cells of the NMJ in a way that may initiate or facilitate the dismantling of the neuromuscular synapse in the presymptomatic stages of ALS. [ABSTRACT FROM AUTHOR]

Mutant ubiquitin UBB+1 is observed in a variety of aging-related neurodegenerative diseases and acts as a potent inhibitor of the ubiquitin proteasome system (UPS). In the present study, we investigated the relationship between impaired UPS (using ectopic expression of UBB+1) and mitochondrial dynamics in astrocytes, which are the most abundant glial cells in the central nervous system. Immunocytochemistry and fluorescence recovery after photobleaching revealed that ectopic expression of UBB+1 induced mitochondrial elongation. We further demonstrated that overexpression of UBB+1 destabilized mitochondrial fission-specific proteins including Drp1, Fis1, and OPA3, but not the mitochondrial fusion-specific proteins Mfn1, Mfn2, and OPA1. The reduction in mitochondrial fission-specific proteins by UBB+1 was prevented by inhibiting the 26 S proteasome using chemical inhibitors, including MG132, lactacystin and epoxomicin. We then assessed the involvement of proteases that target mitochondrial proteins by using various protease inhibitors. Finally, we confirmed that either overexpression of UBB+1 or inhibiting the proteasome can protect astrocytic cells from H2O2-induced cell death compared with control cells. Our results suggest that UBB+1 destabilizes mitochondrial fission-specific proteins, leading to mitochondrial fusion and the subsequent resistance to oxidative stress. We therefore propose a protective role of UBB+1 overexpression or the proteasome inhibition in astrocytes in degenerative brains. [ABSTRACT FROM AUTHOR]

The ubiquitin-proteasome system (UPS) controls intracellular protein turnover in a substrate-specific manner via E3-type ubiquitin ligases. Mammalian fertilization and particularly sperm penetration through the oocyte vitelline coat, the zona pellucida (ZP), is regulated by UPS. We use an extrinsic substrate of the proteasome-dependent ubiquitin-fusion degradation pathway, the mutant ubiquitin UBB, to provide evidence that an E3-type ligase activity exists in sperm-acrosomal fractions. Protein electrophoresis gels from such de novo ubiquitination experiments contained a unique protein band identified by tandem mass spectrometry as being similar to ubiquitin ligase UBR7 (alternative name: C14ORF130). Corresponding mRNA was amplified from boar testis and several variants of the UBR7 protein were detected in boar, mouse and human sperm extracts by Western blotting. Genomic analysis indicated a high degree of evolutionary conservation, remarkably constant purifying selection and conserved testis expression of the UBR7 gene. By immunofluorescence, UBR7 was localized to the spermatid acrosomal cap and sperm acrosome, in addition to hotspots of proteasomal activity in spermatids, such as the cytoplasmic lobe, caudal manchette, nucleus and centrosome. During fertilization, UBR7 remained with the ZP-bound acrosomal shroud following acrosomal exocytosis. Thus, UBR7 is present in the acrosomal cap of round spermatids and within the acrosomal matrix of mature boar spermatozoa. These data provide the first evidence of ubiquitin ligase activity in mammalian spermatozoa and indicate UBR7 involvement in spermiogenesis. [ABSTRACT FROM AUTHOR]

Amyloid-beta (Abeta) peptide is the original causative factor of Alzheimer's disease (AD) according to the amyloid cascade hypothesis. The ubiquitin-proteasome system (UPS), the major intracellular protein quality control system in eukaryotic cells, is related to AD pathogenesis. There is growing evidence showing that there is a tight relationship between Abeta and UPS and this relationship plays an important role in AD pathogenesis. This article reviews the relationship between Abeta and the UPS in terms of the following three aspects: the interaction of the two factors, the ubiquitinating process of Abeta, and impact of dysfunctional UPS on Abeta production. The impairment in the UPS in AD could affect the degradation of Abeta and lead to an abnormal accumulation of Abeta. At the same time, Abeta inhibits the proteasomal activity and subsequently leads to impairment of multivesicular bodies (MVB) sorting pathway, forming an interacting relationship between Abeta and UPS. Mutant ubiquitin (Ub) and ubiquitin-like (UBL) ubiquilin-1 are related to Abeta accumulation. Meanwhile E2 conjugating enzymes, E3 ligases, and de-ubiquitinating enzymes, all of which function in the ubiquitination process, play a pivotal role in the proteasomal degradation of Abeta. Ubiquitin-proteasome system has an immense impact on the amyloidogenic pathway of amyloid precursor protein (APP) processing that generates Abeta. Upregulation in proteasomal degradation of BACE1 and components of gamma-secretase leads to decreased Abeta accumulation. A deep look into the mechanism underlying the interplay between Abeta and UPS may provide alternative therapeutic targets and lead to new drugs and therapies. [ABSTRACT FROM AUTHOR]

Over the last decades, studies from our laboratory and other groups using animal models have shown that iron overload, resulting in iron accumulation in the brain, produces significant cognitive deficits. Iron accumulation in the hippocampus and the basal ganglia has been related to impairments in spatial memory, aversive memory, and recognition memory in rodents. These results are corroborated by studies showing that the administration of iron chelators attenuates cognitive deficits in a variety of animal models of cognitive dysfunction, including aging and Alzheimer's disease models. Remarkably, recent human studies using magnetic resonance image techniques have also shown a consistent correlation between cognitive dysfunction and iron deposition, mostly in the hippocampus, cortical areas, and basal ganglia. These findings may have relevant implications in the light of the knowledge that iron accumulates in brain regions of patients suffering from neurodegenerative diseases. A better understanding of the functional consequences of iron dysregulation in aging and neurological diseases may help to identify novel targets for treating memory problems that afflict a growing aging population. [ABSTRACT FROM AUTHOR]