Wiep Scheper

The unfolded protein response in neurodegenerative diseases: a neuropathological perspective

The unfolded protein response (UPR) is a stress response of the endoplasmic reticulum (ER) to a disturbance in protein folding. The so-called ER stress sensors PERK, IRE1 and ATF6 play a central role in the initiation and regulation of the UPR. The accumulation of misfolded and aggregated proteins is a common characteristic of neurodegenerative diseases. With the discovery of the basic machinery of the UPR, the idea was born that the UPR or part of its machinery could be involved in neurodegenerative diseases like Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis and prion disease. Over the last decade, the UPR has been addressed in an increasing number of studies on neurodegeneration. The involvement of the UPR has been investigated in human neuropathology across different neurological diseases, as well as in cell and mouse models for neurodegeneration. Studies using different disease models display discrepancies on the role and function of the UPR during neurodegeneration, which can often be attributed to differences in methodology. In this review, we will address the importance of investigation of human brain material for the interpretation of the role of the UPR in neurological diseases. We will discuss evidence for UPR activation in neurodegenerative diseases, and the methodology to study UPR activation and its connection to brain pathology will be addressed. More recently, the UPR is recognized as a target for drug therapy for treatment and prevention of neurodegeneration, by inhibiting the function of specific mediators of the UPR. Several preclinical studies have shown a proof-of-concept for this approach targeting the machinery of UPR, in particular the PERK pathway, in different models for neurodegeneration and have yielded paradoxical results. The promises held by these observations will need further support by clarification of the observed differences between disease models, as well as increased insight obtained from human neuropathology.

Endoplasmic reticulum stress activates autophagy but not the proteasome in neuronal cells: implications for Alzheimer's disease

Protein folding stress in the endoplasmic reticulum (ER) may lead to activation of the unfolded protein response (UPR), aimed to restore cellular homeostasis via transcriptional and post-transcriptional mechanisms. ER stress is also reported to activate the ER overload response (EOR), which activates transcription via NF-κB. We previously demonstrated that UPR activation is an early event in pre-tangle neurons in Alzheimers disease (AD) brain. Misfolded and unfolded proteins are degraded via the ubiquitin proteasome system (UPS) or autophagy. UPR activation is found in AD neurons displaying both early UPS pathology and autophagic pathology. Here we investigate whether activation of the UPR and/or EOR is employed to enhance the proteolytic capacity of neuronal cells. Expression of the immunoproteasome subunits β2i and β5i is increased in AD brain. However, expression of the proteasome subunits is not increased by the UPR or EOR. UPR activation does not relocalize the proteasome or increase overall proteasome activity. Therefore proteasomal degradation is not increased by ER stress. In contrast, UPR activation enhances autophagy and LC3 levels are increased in neurons displaying UPR activation in AD brain. Our data suggest that autophagy is the major degradational pathway following UPR activation in neuronal cells and indicate a connection between UPR activation and autophagic pathology in AD brain.

Removing protein aggregates: the role of proteolysis in neurodegeneration.

A common characteristic of neurodegenerative diseases like Alzheimers disease (AD), Parkinsons disease (PD) and Huntingtons disease (HD) is the accumulation of protein aggregates. This reflects a severe disturbance of protein homeostasis, the proteostasis. Here, we review the involvement of the two major proteolytic machineries, the ubiquitin proteasome system (UPS) and the autophagy/lysosomal system, in the pathogenesis of neurodegenerative diseases. These proteolytic systems cooperate to maintain the proteostasis, as is indicated by intricate cross talk. In addition, the UPS and autophagy are regulated by stress pathways that are activated by disturbed proteostasis, like the unfolded protein response (UPR). We will specifically discuss how these proteolytic pathways are affected in neurodegenerative diseases. We will show that there is a differential involvement of the UPS and autophagy in different neurodegenerative disorders. In addition, the proteolytic impairment may be primary or secondary to the pathology. These differences have important implications for the design of therapeutic strategies. The opportunities and caveats of targeting the UPS and autophagy/lysosomal system as a therapeutic strategy in neurodegeneration will be discussed.

The unfolded protein response mediates reversible tau phosphorylation induced by metabolic stress

The unfolded protein response (UPR) is activated in neurodegenerative tauopathies such as Alzheimer’s disease (AD) in close connection with early stages of tau pathology. Metabolic disturbances are strongly associated with increased risk for AD and are a potent inducer of the UPR. Here, we demonstrate that metabolic stress induces the phosphorylation of endogenous tau via activation of the UPR. Strikingly, upon restoration of the metabolic homeostasis, not only the levels of the UPR markers pPERK, pIRE1α and BiP, but also tau phosphorylation are reversed both in cell models as well as in torpor, a physiological hypometabolic model in vivo. Intervention in the UPR using the global UPR inhibitor TUDCA or a specific small-molecule inhibitor of the PERK signaling pathway, inhibits the metabolic stress-induced phosphorylation of tau. These data support a role for UPR-mediated tau phosphorylation as part of an adaptive response to metabolic stress. Failure to restore the metabolic homeostasis will lead to prolonged UPR activation and tau phosphorylation, and may thus contribute to AD pathogenesis. We demonstrate that the UPR is functionally involved in the early stages of tau pathology. Our data indicate that targeting of the UPR may be employed for early intervention in tau-related neurodegenerative diseases.

Rab6 is increased in Alzheimer's disease brain and correlates with endoplasmic reticulum stress

Alzheimers disease (AD) is characterized by deposits of aggregated proteins. Accumulation of aggregation‐prone proteins activates protein quality control mechanisms, such as the unfolded protein response (UPR) in the endoplasmic reticulum (ER). We previously reported upregulation of the UPR marker BiP in AD brain. In this study, we investigated the small GTPase Rab6, which is involved in retrograde Golgi–ER trafficking and may function as a post‐ER quality control system. Using immunohistochemistry and semiquantitative Western blotting, the expression of Rab6 was analysed in hippocampus, entorhinal and temporal cortex of 10u2003AD patients and six nondemented control subjects. Rab6 is upregulated in AD temporal cortex from Braak stage 3/4, the same stage that UPR activation is found. We observe increased neuronal Rab6 immunoreactivity in all brain areas examined. Although some neurones show colocalization of immunoreactivity for Rab6 and hyperphosphorylated tau, strong Rab6 staining does not colocalize with tangles. We find a highly significant correlation between the Rab6 and BiP levels. In vitro data show that Rab6 is not upregulated as a result of UPR activation or proteasome inhibition indicating an independent regulatory mechanism. Our data suggest that ER and post‐ER protein quality control mechanisms are activated early in the pathology of AD.

Intracellular accumulation of aggregated pyroglutamate amyloid beta: convergence of aging and Aβ pathology at the lysosome

Deposition of aggregated amyloid beta (Aβ) is a major hallmark of Alzheimer’s disease (AD)—a common age-related neurodegenerative disorder. Typically, Aβ is generated as a peptide of varying lengths. However, a major fraction of Aβ peptides in the brains of AD patients has undergone posttranslational modifications, which often radically change the properties of the peptides. Aβ3(pE)-42 is an N-truncated, pyroglutamate-modified variant that is abundantly present in AD brain and was suggested to play a role early in the pathogenesis. Here we show that intracellular accumulation of oligomeric aggregates of Aβ3(pE)-42 results in loss of lysosomal integrity. Using a novel antibody specific for aggregates of AβpE3, we show that in postmortem human brain tissue, aggregated AβpE3 is predominantly found in the lysosomes of both neurons and glial cells. Our data further demonstrate that AβpE3 is relatively resistant to lysosomal degradation, which may explain its accumulation in the lysosomes. The intracellular AβpE3 aggregates increase in an age-dependent manner. The results presented in this study support a model where Aβ pathology and aging converge, leading to accumulation of the degradation-resistant pE-modified Aβ in the lysosomes, lysosomal dysfunction, and neurodegeneration.

From alpha to omega with Abeta: targeting the multiple molecular appearances of the pathogenic peptide in Alzheimer's disease.

Amyloid beta (Abeta) is the main component of one of the major pathological hallmarks of Alzheimers disease and is generally considered as one of the earliest factors that induce the pathogenic cascade. Abeta is produced from a larger precursor protein through proteolytic cleavage by secretase activities, which results in fragments that differ in size depending on the cleavage site used to create the C-terminus. In addition, heterogeneity at the N-terminus is created by proteases/peptidases. Moreover, various amino acid modifications further enhance the heterogeneity of Abeta that accumulates in Alzheimer brain. All these species with their different N-and C termini, with or without modifications have different aggregation properties. Abeta requires an aggregated state to be pathogenic and the exact aggregation state is a major determinant of the cellular effects of Abeta: smaller oligomeric aggregates are more neurotoxic, whereas large fibrillar aggregates are generally more associated with a glial response. It is therefore increasingly clear that Abeta is not a single entity, but a peptide with multiple molecular appearances. In this review we will discuss the mechanisms leading to the generation of the different Abeta species and their involvement in Alzheimer pathogenesis. This will be discussed in the framework of therapeutic approaches that target one of the steps in the biogenesis of toxic Abeta species: inhibition of the formation of Abeta, inhibition of aggregation and stimulation of its degradation or clearance.

Alternative splicing in the N-terminus of Alzheimer’s presenilin 1

Abstract.Presenilin 1 (PS1) is mutated in the majority of familial cases of Alzheimer disease (AD). Although it is clear that PS1 is involved in the processing of the amyloid precursor protein (APP), the exact function of PS1 is still elusive. Human presenilin 1 (PS1) is alternatively spliced, resulting in the presence or absence of a four-amino acid motif, VRSQ, in the PS1 N-terminus. In human tissues, both isoforms are expressed. Here we report that mouse and rat only express the longer PS1 isoform. The presence of this motif introduces a potential phosphorylation site for protein kinase C. Because the splice occurs in the region of PS1 that we have previously shown to bind to rabGDI, this might provide a regulatory mechanism for this interaction. Our data show that the –VRSQ isoform binds rabGDI, but the +VRSQ does not. Moreover, mutation of the putatively phosphorylated threonine in PS1 disrupts the binding to rabGDI, showing its importance for the interaction. To our knowledge this is the first study showing a functional difference between PS1 splice variants. The possible consequences for APP processing and the pathogenesis of AD are discussed.

Nanoparticles against Alzheimer's disease: PEG-PACA nanoparticles are able to link the aβ-peptide and influence its aggregation kinetic.

D. Brambilla1, R. Verpillot2, L. De Kimpe3, M. Taverna2, B. Le Droumaguet1, J. Nicolas1, M. Canovi4, M. Gobbi4, M. Salmona4, V. Nicolas5, W. Scheper3, P. Couvreur1, K. Andrieux1 1 Laboratory of Physical-Chemistry, Pharmaceutical Technology and Biopharmacy, UMR CNRS 8612, University of Paris-Sud 11, Faculty of Pharmacy, 5 Rue J-B Clement, 92296 Chatenay-Malabry, France; 2 Laboratory of Proteins and Nanotechnology in Separation Science (LPNSS), UMR CNRS-8612, University of Paris-Sud 11, Faculty of Pharmacy, 5 Rue J-B Clement, 92296 Chatenay-Malabry, France; 3 Neurogenetics Laboratory, Academic Medical Center, Amsterdam, The Netherlands 4 Istituto di Ricerche Farmacologiche “Mario Negri”, Milano, Italy; 5 Therapeutical Innovation Institute (IFR141 ITFM), University of Paris-Sud 11, Faculty of Pharmacy, 5 Rue J-B Clement, 92296 Chatenay-Malabry, France;

Inhibition of Endoplasmic Reticulum Associated Degradation Reduces Endoplasmic Reticulum Stress and Alters Lysosomal Morphology and Distribution

Disturbances in proteostasis are observed in many neurodegenerative diseases. This leads to activation of protein quality control to restore proteostasis, with a key role for the removal of aberrant proteins by proteolysis. The unfolded protein response (UPR) is a protein quality control mechanism of the endoplasmic reticulum (ER) that is activated in several neurodegenerative diseases. Recently we showed that the major proteolytic pathway during UPR activation is via the autophagy/lysosomal system. Here we investigate UPR induction if the other major proteolytic pathway of the ER -ER associated degradation (ERAD)-is inhibited. Surprisingly, impairment of ERAD results in decreased UPR activation and protects against ER stress toxicity. Autophagy induction is not affected under these conditions, however, a striking relocalization of the lysosomes is observed. Our data suggest that a protective UPR-modulating mechanism is activated if ERAD is inhibited, which involves lysosomes. Our data provide insight in the cross-talk between proteolytic pathways involved in ER proteostasis. This has implications for neurodegenerative diseases like Alzheimer’s disease where disturbed ER proteostasis and proteolytic impairment are early phenomena in the pathology.