Removal Of Protein Aggregates Research Articles

Role of TFEB in Huntington's Disease.

Huntington's disease (HD) is an autosomal dominant neurodegenerative disease caused by an expansion of the CAG trinucleotide repeat in exon 1 of the huntingtin (HTT) gene. This expansion leads to a polyglutamine (polyQ) tract at the N-terminal end of HTT, which reduces the solubility of the protein and promotes its accumulation. Inefficient clearance of mutant HTT (mHTT) by the proteasome or autophagy-lysosomal system leads to accumulation of oligomers and toxic protein aggregates in neurons, resulting in impaired proteolytic systems, transcriptional dysregulation, impaired axonal transport, mitochondrial dysfunction and cellular energy imbalance. Growing evidence suggests that the accumulation of mHTT aggregates and autophagic and/or lysosomal dysfunction are the major pathogenic mechanisms underlying HD. In this context, enhancing autophagy may be an effective therapeutic strategy to remove protein aggregates and improve cell function. Transcription factor EB (TFEB), a master transcriptional regulator of autophagy, controls the expression of genes critical for autophagosome formation, lysosomal biogenesis, lysosomal function and autophagic flux. Consequently, the induction of TFEB activity to promote intracellular clearance may be a therapeutic strategy for HD. However, while some studies have shown that overexpression of TFEB facilitates the clearance of mHTT aggregates and ameliorates the disease phenotype, others indicate such overexpression may lead to mHTT co-aggregation and worsen disease progression. Further studies are necessary to confirm whether TFEB modulation could be an effective therapeutic strategy against mHTT-mediated toxicity in different disease models.

Traditional Chinese medicine and its active substances reduce vascular injury in diabetes via regulating autophagic activity.

Due to its high prevalence, poor prognosis, and heavy burden on healthcare costs, diabetic vascular complications have become a significant public health issue. Currently, the molecular and pathophysiological mechanisms underlying diabetes-induced vascular complications remain incompletely understood. Autophagy, a highly conserved process of lysosomal degradation, maintains intracellular homeostasis and energy balance via removing protein aggregates, damaged organelles, and exogenous pathogens. Increasing evidence suggests that dysregulated autophagy may contribute to vascular abnormalities in various types of blood vessels, including both microvessels and large vessels, under diabetic conditions. Traditional Chinese medicine (TCM) possesses the characteristics of "multiple components, multiple targets and multiple pathways," and its safety has been demonstrated, particularly with minimal toxicity in liver and kidney. Thus, TCM has gained increasing attention from researchers. Moreover, recent studies have indicated that Chinese herbal medicine and its active compounds can improve vascular damage in diabetes by regulating autophagy. Based on this background, this review summarizes the classification, occurrence process, and related molecular mechanisms of autophagy, with a focus on discussing the role of autophagy in diabetic vascular damage and the protective effects of TCM and its active compounds through the regulation of autophagy in diabetes. Moreover, we systematically elucidate the autophagic mechanisms by which TCM formulations, individual herbal extracts, and active compounds regulate diabetic vascular damage, thereby providing new candidate drugs for clinical treatment of vascular complications in diabetes. Therefore, further exploration of TCM and its active compounds with autophagy-regulating effects holds significant research value for achieving targeted therapeutic approaches for diabetic vascular complications.

Preserved Autophagy in Cognitive Intact Non‐Demented Individuals with Alzheimer’s Neuropathology

AbstractBackgroundAlzheimer’s disease (AD) is a terminal neurodegenerative disease and the most common cause of dementia. Notably, some individuals, here referred to as Non‐Demented individuals with Alzheimer’s Neuropathology (NDAN) are cognitively intact despite the presence of neuropathology consistent with fully symptomatic AD and have decreased Tau oligomers at synapses. Growing evidence supports that dysfunctional autophagy, the primary cell mechanism responsible for removing protein aggregates and a route of clearance for Tau in healthy neurons, is a major finding in demented AD patients. However, the association of autophagy with maintenance of cognitive integrity in resilient individuals has not been evaluated. Thus, the understanding of the neuroprotective mechanisms to evade cognitive decline to preserve synaptic functional integrity is important for the development of therapeutic approaches to prevent dementia.MethodHere we evaluated autophagy as a key protective mechanism for maintenance of cognitive integrity in NDAN subjects by efficient removal of tau oligomers. Using postmortem hippocampal samples from age‐matched healthy controls (n = 6), AD (n = 7) and NDAN (n = 7) we performed immunofluorescence and comparative western blot analyses for regulatory molecules and ATGs (autophagy‐related genes) involved in autophagy. Tauopathy was accessed by the expression and phosphorylation levels at Ser202/Thr205 (AT8), Thr231 (AT180) and Ser396 (PHF13.6) of Tau oligomers.ResultWe found that NDAN subjects have significantly increased regulatory Beclin‐1, and Atg16L, Atg12, Atg3, Atg5, Atg7, LC3AB, and proteasomal PSMA5 but not lysosomal LAMP1 as compared to AD. Expression levels were comparable with healthy controls. Autophagic proteins, except p62, were negatively correlated to Tau oligomer (>65 kDa), but not to total Tau (>45 kDa), indicating autophagy being specific to tau oligomer clearance. Tau oligomer expression and levels of phosphorylation at Ser202/Thr205, Thr231, and Ser396 were significantly reduced in hippocampal tissue in NDAN as compared to AD, also confirmed by immunofluorescence analyses of Tau tangles.ConclusionOur results indicate, for the first time, intact autophagy and associated reduced tauopathy as one of protective mechanism in cognitive intact NDAN. This novel observation supports the potential of autophagy‐induced strategies in AD therapeutics

Emulsification Properties of Garlic Aqueous Extract: Effect of Heat Treatment and pH Modification.

Despite the broad research available in the literature dealing with garlic health benefits, little information is found regarding the functional properties of garlic components. The aim of this study was to evaluate the emulsification properties of garlic water-soluble compounds (GWSC), encompassing proteins, saponins, and carbohydrates, after heat treatment (10 min at 95 °C) or pH adjustments (2.5, 3.5, and 7.8). After the various treatments, the extracts were used as such or filtrated (0.45 µm), and 10% soybean oil-in-water emulsions were prepared using low (0.48%) or high (6.55% wt/wt) extract concentrations. Results showed that whereas at low GWSC concentrations, both heating and acidifying resulted in the formation of bigger oil droplet sizes (i.e., from d32 = 0.36 µm using unmodified extract to d32 = 7-22 µm at pH 2.5 with or without extract filtration), the effects were opposite at the highest GWSC concentration. In the latter, heat treatment clearly reduced the droplet size as observed from the micrographs as well as the degree of creaming, though the occurrence of depletion and/or bridging flocculation was still strong. The acidification of the extract at this high GWSC concentration significantly reduced the droplet size, as observed from the micrographs; however, a strong flocculation was observed. Removal of protein aggregates, and possibly also saponin micelles, from the extract resulted in an obvious increase in emulsion droplet size. This research brings valuable insights on this study and utilisation of novel natural food emulsifiers from plant sources.

Oxidative Stress and Cellular Protein Accumulation Are Present in Keratoconus, Macular Corneal Dystrophy, and Fuchs Endothelial Corneal Dystrophy

The aim of the study was to investigate oxidative stress as well as cellular protein accumulation in corneal diseases including keratoconus (KC), macular corneal dystrophy (MCD), and Fuchs endothelial corneal dystrophy (FECD) at their primary affecting sites. Corneal buttons from KC, MCD, and FECD patients, as well as healthy controls, were analyzed immunohistochemically to evaluate the presence of oxidative stress and the function of the proteostasis network. 4-Fydroxynonenal (4-HNE) was used as a marker of oxidative stress, whereas the levels of catalase and heat-shock protein 70 (HSP70) were analyzed to evaluate the response of the antioxidant defense system and molecular chaperones, respectively. Sequestosome 1 (SQSTM1) levels were determined to assess protein aggregation and the functionality of autophagic degradation. Basal epithelial cells of the KC samples showed increased levels of oxidative stress marker 4-HNE and antioxidant enzyme catalase together with elevated levels of HSP70 and accumulation of SQSTM1. Corneal stromal cells and endothelial cells from MCD and FECD samples, respectively, showed similarly increased levels of these markers. All corneal diseases showed the presence of oxidative stress and activation of the molecular chaperone response to sustain protein homeostasis. However, the accumulation of protein aggregates suggests insufficient function of the protective mechanisms to limit the oxidative damage and removal of protein aggregates via autophagy. These results suggest that oxidative stress has a role in KC, MCD, and FECD at the cellular level as a secondary outcome. Thus, antioxidant- and autophagy-targeted therapies could be included as supporting care when treating KC or corneal dystrophies.

Preserved autophagy in cognitively intact non-demented individuals with Alzheimer's neuropathology.

Growing evidence supports that dysfunctional autophagy, the major cell mechanism responsible for removing protein aggregates and a route of clearance for Tau in healthy neurons, is a major finding in demented Alzheimer's disease (AD) patients. However, the association of autophagy with maintenance of cognitive integrity in resilient individuals who have AD neuropathology but remain non-demented (NDAN) has not been evaluated. Using post mortem brain samples from age-matched healthy control, AD, and NDAN subjects, we evaluated autophagy in relation to Tau pathology using Western blot, immunofluorescence and RNA-seq. Compared to AD patients, NDAN subjects had preserved autophagy and reduced tauopathy. Furthermore, expression of autophagy genes and AD-related proteins were significantly associated in NDAN compared to AD and control subjects. Our results suggest preserved autophagy is a protective mechanism that maintains cognitive integrity in NDAN individuals. This novel observation supports the potential of autophagy-inducing strategies in AD therapeutics. NDAN subjects have preserved autophagic protein levels comparable with control subjects. Compared to control subjects, NDAN subjects have significantly reduced Tau oligomers and PHF Tau phosphorylation at synapses that negatively correlate with autophagy markers. Transcription of autophagy genes strongly associates with AD-related proteins in NDAN donors.

Increase of autophagy marker p62 in the placenta from pregnant women with preeclampsia

Preeclampsia (PE) is a multisystemic disorder characterized by abnormal placentation. Autophagy is a lysosomal degradation pathway that removes protein aggregates and damaged organelles, and it seems to be essential for cell survival during stress, hypoxia, and for implantation and development of the placenta. p62/SQSTM1 is an autophagy marker that not only binds proteins destined for elimination but is also constitutively degraded by this mechanism. Considering that the placenta plays an important role in the pathogenesis of PE, the present study aimed to evaluate the gene and protein expression of p62/SQSTM1 in placentas from pregnant women with PE. Placental tissues from 20 women with PE classified into three groups according to gestational age, 27–31 weeks (n = 8); 32–36 weeks (n = 6); 37–39 weeks (n = 6), and 20 normotensives (NT) pregnant women were collected and employed for p62/SQSTM1 expression by quantitative polymerase chain reaction (qPCR), immunohistochemistry and enzyme-linked immunosorbent assay (ELISA) techniques. p62/SQSTM1 mRNA levels were significantly lower, while protein expression was significantly higher in the placenta of pregnant women with PE than in NT pregnant women, and these results remained similar after separating the groups by gestational age. In conclusion, the results suggest that there is a reduction of autophagic activity in pregnant women with PE. Studies involving cross-talk between autophagy, inflammasomes, nuclear transcription factor (NF-κB) activation pathways, and aggregation of protein in the placenta from women with PE might help to better understand the pathogenesis of this important obstetric pathology.

Down-regulation of autophagy proteins is associated with higher mTOR expression in the placenta of pregnant women with preeclampsia.

Autophagy is a lysosomal degradation pathway that removes protein aggregates and damaged organelles maintaining cellular integrity. It seems to be essential for cell survival during stress, starvation, hypoxia, and consequently to the placenta implantation and development. Preeclampsia (PE) is a multisystemic disorder characterized by the onset of hypertension associated or not with proteinuria and other maternal complications. Considering that the placenta seems to play an important role in the pathogenesis of PE, the objective of the present study was to evaluate protein levels of light chain protein (LC3), beclin-1, and the mammalian target of rapamycin (mTOR) in the placenta of pregnant women with PE. Placental tissues collected from 20 women with PE and 20 normotensive (NT) pregnant women were evaluated for LC3, beclin-1, and mTOR expression by qPCR and immunohistochemistry. The mRNA for LC3 and beclin-1 were significantly lower, while mTOR gene expression was significantly higher in the placenta of pregnant women with PE than in the NT group. Placentas of PE women showed significantly decreased protein expression of LC3-II and beclin-1, whereas mTOR was significantly increased compared with the NT pregnant women. There was a negative correlation between protein expression of mTOR and LC3-II in the placental tissue of PE women. In conclusion, the results showed autophagy deficiency suggesting that failure in this degradation process may contribute to the pathogenesis of PE; however, new studies involving cross-talk between autophagy and inflammatory molecular mechanisms might help to better understand the autophagy process in this obstetric pathology.

Autophagy in Cisplatin Nephrotoxicity during Cancer Therapy.

Simple SummaryCisplatin is a broadly used chemotherapy drug, but its use and efficacy are limited by its nephrotoxicity. Autophagy protects against kidney injury during cisplatin exposure but may reduce the efficacy of chemotherapy by protecting cancer cells. In this review, we describe the role and regulation of autophagy in cisplatin-induced nephrotoxicity and discuss the therapeutic advances and challenges of targeting autophagy in chemotherapy.Cisplatin is a widely used chemotherapeutic agent but its clinical use is often limited by nephrotoxicity. Autophagy is a lysosomal degradation pathway that removes protein aggregates and damaged or dysfunctional cellular organelles for maintaining cell homeostasis. Upon cisplatin exposure, autophagy is rapidly activated in renal tubule cells to protect against acute cisplatin nephrotoxicity. Mechanistically, the protective effect is mainly related to the clearance of damaged mitochondria via mitophagy. The role and regulation of autophagy in chronic kidney problems after cisplatin treatment are currently unclear, despite the significance of research in this area. In cancers, autophagy may prevent tumorigenesis, but autophagy may reduce the efficacy of chemotherapy by protecting cancer cells. Future research should focus on developing drugs that enhance the anti-tumor effects of cisplatin while protecting kidneys during cisplatin chemotherapy.

Targeting autophagy in neurodegenerative diseases: From molecular mechanisms to clinical therapeutics.

Many neurodegenerative diseases are associated with pathological aggregation of proteins in neurons. Autophagy is a natural self-cannibalization process that can act as a powerful mechanism to remove aged and damaged organelles as well as protein aggregates. It has been shown that promoting autophagy can attenuate or delay neurodegeneration by removing protein aggregates. In this paper, we will review the role of autophagy in Alzheimer's disease (AD), Parkinson's Disease (PD), and Huntington's Disease (HD) and discuss opportunities and challenges of targeting autophagy as a potential therapeutic avenue for treatment of these common neurodegenerative diseases.

Friend or Foe: Lipid Droplets as Organelles for Protein and Lipid Storage in Cellular Stress Response, Aging and Disease.

Lipid droplets (LDs) were considered as a mere lipid storage organelle for a long time. Recent evidence suggests that LDs are in fact distinct and dynamic organelles with a specialized proteome and functions in many cellular roles. As such, LDs contribute to cellular signaling, protein and lipid homeostasis, metabolic diseases and inflammation. In line with the multitude of functions, LDs interact with many cellular organelles including mitochondria, peroxisomes, lysosomes, the endoplasmic reticulum and the nucleus. LDs are highly mobile and dynamic organelles and impaired motility disrupts the interaction with other organelles. The reduction of interorganelle contacts results in a multitude of pathophysiologies and frequently in neurodegenerative diseases. Contacts not only supply lipids for β-oxidation in mitochondria and peroxisomes, but also may include the transfer of toxic lipids as well as misfolded and harmful proteins to LDs. Furthermore, LDs assist in the removal of protein aggregates when severe proteotoxic stress overwhelms the proteasomal system. During imbalance of cellular lipid homeostasis, LDs also support cellular detoxification. Fine-tuning of LD function is of crucial importance and many diseases are associated with dysfunctional LDs. We summarize the current understanding of LDs and their interactions with organelles, providing a storage site for harmful proteins and lipids during cellular stress, aging inflammation and various disease states.

Metformin rescues muscle function in BAG3 myofibrillar myopathy models

ABSTRACT Dominant de novo mutations in the co-chaperone BAG3 cause a severe form of myofibrillar myopathy, exhibiting progressive muscle weakness, muscle structural failure, and protein aggregation. To elucidate the mechanism of disease in, and identify therapies for, BAG3 myofibrillar myopathy, we generated two zebrafish models, one conditionally expressing BAG3P209L and one with a nonsense mutation in bag3. While transgenic BAG3P209L-expressing fish display protein aggregation, modeling the early phase of the disease, bag3 -/- fish exhibit exercise dependent fiber disintegration, and reduced swimming activity, consistent with later stages of the disease. Detailed characterization of the bag3 -/- fish, revealed an impairment in macroautophagic/autophagic activity, a defect we confirmed in BAG3 patient samples. Taken together, our data highlights that while BAG3P209L expression is sufficient to promote protein aggregation, it is the loss of BAG3 due to its sequestration within aggregates, which results in impaired autophagic activity, and subsequent muscle weakness. We therefore screened autophagy-promoting compounds for their effectiveness at removing protein aggregates, identifying nine including metformin. Further evaluation demonstrated metformin is not only able to bring about the removal of protein aggregates in zebrafish and human myoblasts but is also able to rescue the fiber disintegration and swimming deficit observed in the bag3 −/- fish. Therefore, repurposing metformin provides a promising therapy for BAG3 myopathy. Abbreviations:ACTN: actinin, alpha; BAG3: BAG cochaperone 3; CRYAB: crystallin alpha B; DES: desmin; DMSO: dimethyl sulfoxide; DNAJB6: DnaJ heat shock protein family (Hsp40) member B6; dpf: days post fertilization; eGFP: enhanced green fluorescent protein; FDA: Food and Drug Administration; FHL1: four and a half LIM domains 1; FLNC: filamin C; hpf: hours post-fertilization; HSPB8: heat shock protein family B [small] member 8; LDB3/ZASP: LIM domain binding 3; MYOT: myotilin; TTN: titin; WT: wild-type.

Mechanism of Activation of Mechanistic Target of Rapamycin Complex 1 by Methionine.

Nutrients are closely involved in the regulation of lifespan and metabolic health. Cellular activities, such as the regulation of metabolism, growth, and aging, are mediated by a network of nutrients and nutrient-sensing pathways. Among the nutrient-sensing pathways, the mechanistic target of rapamycin complex 1 (mTORC1) acts as the central regulator of cellular functions, which include autophagy. Autophagy plays a significant role in the removal of protein aggregates and damaged or excess organelles, including mitochondria, to maintain intracellular homeostasis, which is involved in lifespan extension and cardiometabolic health. Moreover, dietary methionine restriction may have a beneficial effect on lifespan extension and metabolic health. In contrast, methionine may activate mTORC1 and suppress autophagy. As the mechanism of methionine sensing on mTORC1, SAMTOR was identified as a sensor of S-adenosyl methionine (SAM), a metabolite of methionine, in the cytoplasm. Conversely, methionine may activate the mTORC1 signaling pathway through the activation of phosphatase 2A (PP2A) because of increased methylation in response to intracellular SAM levels. In this review, we summarized the recent findings regarding the mechanism via which methionine activates mTORC1.

Non-apoptotic roles of apoptotic proteases: new tricks for an old dog.

This Open Question article highlights current advances in the study of non-apoptotic roles of apoptotic proteins. Apoptosis is a highly regulated and energy-requiring process in which cells actively kill themselves. Apoptosis helps remove extra cells to sculpt organs during embryo development and culls damaged cells throughout the body. Apoptosis relies on evolutionarily conserved proteins that include a family of proteases called caspases. Caspases activity has long been considered a hallmark of apoptosis. Yet an emerging body of literature indicates that caspase activity is required for a number of non-lethal processes that range from sculpting cells, removing protein aggregates, changing cell identity during differentiation or de-differentiation, and rebuilding tissues. Failure in each of these processes is associated with human disease. This article is not meant to be an exhaustive review but an introduction to the subject for an educated public, with caspases as a gateway example. I propose that it is time to explore non-apoptotic roles of caspases and other apoptotic proteins, in order to better understand their non-apoptosis function and to leverage new knowledge into new therapies.

Drosophila NUAK functions with Starvin/BAG3 in autophagic protein turnover.

The inability to remove protein aggregates in post-mitotic cells such as muscles or neurons is a cellular hallmark of aging cells and is a key factor in the initiation and progression of protein misfolding diseases. While protein aggregate disorders share common features, the molecular level events that culminate in abnormal protein accumulation cannot be explained by a single mechanism. Here we show that loss of the serine/threonine kinase NUAK causes cellular degeneration resulting from the incomplete clearance of protein aggregates in Drosophila larval muscles. In NUAK mutant muscles, regions that lack the myofibrillar proteins F-actin and Myosin heavy chain (MHC) instead contain damaged organelles and the accumulation of select proteins, including Filamin (Fil) and CryAB. NUAK biochemically and genetically interacts with Drosophila Starvin (Stv), the ortholog of mammalian Bcl-2-associated athanogene 3 (BAG3). Consistent with a known role for the co-chaperone BAG3 and the Heat shock cognate 71 kDa (HSC70)/HSPA8 ATPase in the autophagic clearance of proteins, RNA interference (RNAi) of Drosophila Stv, Hsc70-4, or autophagy-related 8a (Atg8a) all exhibit muscle degeneration and muscle contraction defects that phenocopy NUAK mutants. We further demonstrate that Fil is a target of NUAK kinase activity and abnormally accumulates upon loss of the BAG3-Hsc70-4 complex. In addition, Ubiquitin (Ub), ref(2)p/p62, and Atg8a are increased in regions of protein aggregation, consistent with a block in autophagy upon loss of NUAK. Collectively, our results establish a novel role for NUAK with the Stv-Hsc70-4 complex in the autophagic clearance of proteins that may eventually lead to treatment options for protein aggregate diseases.

A Diversity of Selective Autophagy Receptors Determines the Specificity of the Autophagy Pathway.

The clearance of surplus, broken, or dangerous components is key for maintaining cellular homeostasis. The failure to remove protein aggregates, damaged organelles, or intracellular pathogens leads to diseases, including neurodegeneration, cancer, and infectious diseases. Autophagy is the evolutionarily conserved pathway that sequesters cytoplasmic components in specialized vesicles, autophagosomes, which transport the cargo to the degradative compartments (vacuoles or lysosomes). Research during the past few decades has elucidated how autophagosomes engulf their substrates selectively. This type of autophagy involves a growing number of selective autophagy receptors (SARs) (e.g., Atg19 in yeasts, p62/SQSTM1 in mammals), which bind to the cargo and simultaneously engage components of the core autophagic machinery via direct interaction with the ubiquitin-like proteins (UBLs) of the Atg8/LC3/GABARAP family and adaptors, Atg11 (in yeasts) or FIP200 (in mammals). In this Review, we critically discuss the biology of the SARs with special emphasis on their interactions with UBLs.

P.77Repurposing of metformin identified as a potential therapy in models of BAG3 myofibrillar myopathy

Dominant de novo mutations in the co-chaperone BAG3 cause a severe form of myofibrillar myopathy, exhibiting progressive muscle weakness, muscle structural failure, and protein aggregation. To identify therapies we generated two zebrafish models, one conditionally expressing BAG3P209L and one with a nonsense mutation in bag3. Whilst transgenic BAG3P209Lexpressing fish display protein aggregation, modelling the early phase of the disease, bag3−/− fish demonstrate impaired autophagic activity, exercise dependent fibre disintegration, and reduced swimming activity, consistent with later stages. Having confirmed the presence of impaired autophagy in patient samples we utilised the zebrafish model to screen a library of autophagy promoting compounds for their effectiveness at removing protein aggregates, identifying nine including Metformin. Further evaluation in our models demonstrated Metformin is not only able to remove the protein aggregates in zebrafish and human myoblasts, but is also able to rescue the fibre disintegration and swimming deficit observed in the bag3−/− fish. Therefore, repurposing Metformin provides a promising therapy for BAG3 myopathy. This study therefore illustrates an approach for the identification of treatments for rare neuromuscular diseases, using genetically matched animal models to identify mechanism, confirmation of this mechanism in patients, a targeted drug screen, and demonstration of efficacy in both an animal and human cell model, providing the necessary data for clinical translation of the findings.

Emerging Role of Genetic Alterations Affecting Exosome Biology in Neurodegenerative Diseases.

The abnormal deposition of proteins in brain tissue is a common feature of neurodegenerative diseases (NDs) often accompanied by the spread of mutated proteins, causing neuronal toxicity. Exosomes play a fundamental role on their releasing in extracellular space after endosomal pathway activation, allowing to remove protein aggregates by lysosomal degradation or their inclusion into multivesicular bodies (MVBs), besides promoting cellular cross-talk. The emerging evidence of pathogenic mutations associated to ND susceptibility, leading to impairment of exosome production and secretion, opens a new perspective on the mechanisms involved in neurodegeneration. Recent findings suggest to investigate the genetic mechanisms regulating the different exosome functions in central nervous system (CNS), to understand their role in the pathogenesis of NDs, addressing the identification of diagnostic and pharmacological targets. This review aims to summarize the mechanisms underlying exosome biogenesis, their molecular composition and functions in CNS, with a specific focus on the recent findings invoking a defective exosome biogenesis as a common biological feature of the major NDs, caused by genetic alterations. Further definition of the consequences of specific genetic mutations on exosome biogenesis and release will improve diagnostic and pharmacological studies in NDs.

Targeting Autophagy for the Treatment of Alzheimer's Disease: Challenges and Opportunities.

Alzheimer’s disease (AD) is the most common type of dementia which characterized by a progressive loss of memory and cognitive function due to degeneration of synapses and axons. Currently, there is no cure for AD. Deposition of extracellular amyloid-β (Aβ) plaques and intracellular tau neurofibrillary tangles (NFTs) are two hallmark pathologic changes in the brains of Alzheimer’s patients. Autophagy is the major mechanism in cells responsible for removing protein aggregates. Accumulation of immature autophagic vacuoles (AVs) in dystrophic neurites of Alzheimer patients’ brains suggests that autophagy process is disrupted. Till now, it is far from clear what role autophagy plays in AD, a causative role, a protective role, or just a consequence of the disease process itself. To design more effective therapeutic strategies towards this devastating disorder, it is essential to understand the exact role of autophagy played during different stages of AD.

Abstract 909: Determining the Role of the Co-chaperone BAG3 at the Cardiac Myofilament

Bcl-2-associated athanogene 3 (BAG3) is an adaptor protein often associated with protein quality control. In the heart, decreased BAG3 expression is linked to dilated cardiomyopathy. Mutations to BAG3 can result in myofibrillar disarray, indicating a possible role at the sarcomere. One previous study found BAG3 has a role in maintaining sarcomere structure, however this was in neonatal myocytes. Whether BAG3 has any role in the myofilament of the adult myocyte is unknown. To determine if BAG3 is present in the cardiac sarcomere and identify the myofilament-BAG3 interactome, we used co-immunoprecipitation of the myofilament fraction from mouse LV and mass spectrometry. This revealed a robust interaction of BAG3 with the myofilament. Furthermore, BAG3 interacted with a heat shock protein (HSPB8) known to assist in the removal of protein aggregates, which are abundant after MI. To determine the mechanistic role of BAG3, we overexpressed the protein in mice hearts using adeno-associated virus 9. At baseline, overexpression caused a slight decrease in skinned myocyte calcium sensitivity compared to WT. This effect may be due to BAG3’s interaction with β-adrenergic receptors, which can cause PKA phosphorylation of thin filament proteins known to reduce calcium sensitivity. Preliminary data support this. We then exposed these mice to left anterior descending coronary artery ligation to induce MI. Compared to shams, MI decreased maximum calcium activated force (F max ) in the WT mice, but BAG3 overexpression restored F max to near sham levels. We also studied the functional impact of a known disease-causing mutation discovered in humans, P209L. Transgenic overexpression of P209L BAG3 had no effect on F max , calcium sensitivity, or cooperativity, suggesting the mutation may not interfere with domains in BAG3 required for its myofilament role. This work is the first to show a functional role for BAG3 in the adult myofilament. However, our findings reveal a complex interaction in which the effect of altered BAG3 expression varies with context. The P209L mutation findings suggest that not all BAG3 mutations have shared phenotypes, a conclusion supported by recent clinical work. Further research is necessary to understand BAG3’s myofilament role in health and disease.