Protein Quality Control System Research Articles

Compartment-specific aggregases direct distinct nuclear and cytoplasmic aggregate deposition.

Disruption of the functional protein balance in living cells activates protective quality control systems to repair damaged proteins or sequester potentially cytotoxic misfolded proteins into aggregates. The established model based on Saccharomyces cerevisiae indicates that aggregating proteins in the cytosol of eukaryotic cells partition between cytosolic juxtanuclear (JUNQ) and peripheral deposits. Substrate ubiquitination acts as the sorting principle determining JUNQ deposition and subsequent degradation. Here, we show that JUNQ unexpectedly resides inside the nucleus, defining a new intranuclear quality control compartment, INQ, for the deposition of both nuclear and cytosolic misfolded proteins, irrespective of ubiquitination. Deposition of misfolded cytosolic proteins at INQ involves chaperone-assisted nuclear import via nuclear pores. The compartment-specific aggregases, Btn2 (nuclear) and Hsp42 (cytosolic), direct protein deposition to nuclear INQ and cytosolic (CytoQ) sites, respectively. Intriguingly, Btn2 is transiently induced by both protein folding stress and DNA replication stress, with DNA surveillance proteins accumulating at INQ. Our data therefore reveal a bipartite, inter-compartmental protein quality control system linked to DNA surveillance via INQ and Btn2.

Oxidative Stress and Protein Quality Control Systems in the Aged Canine Brain as a Model for Human Neurodegenerative Disorders

Aged dogs are considered the most suitable spontaneous animal model for studying normal aging and neurodegenerative diseases. Elderly canines naturally develop cognitive dysfunction and neuropathological hallmarks similar to those seen in humans, especially Alzheimer's disease-like pathology. Pet dogs also share similar living conditions and diets to humans. Oxidative damage accumulates in the canine brain during aging, making dogs a valid model for translational antioxidant treatment/prevention studies. Evidence suggests the presence of detective protein quality control systems, involving ubiquitin-proteasome system (UPS) and Heat Shock Proteins (HSPs), in the aged canine brain. Further studies on the canine model are needed to clarify the role of age-related changes in UPS activity and HSP expression in neurodegeneration in order to design novel treatment strategies, such as HSP-based therapies, aimed at improving chaperone defences against proteotoxic stress affecting brain during aging.

Endoplasmic Reticulum Stress Response and Mutant Protein Degradation in CHO Cells Accumulating Antithrombin (C95R) in Russell Bodies.

Newly synthesized secretory proteins are folded and assembled in the endoplasmic reticulum (ER), where an efficient protein quality control system performs a critically important function. When unfolded or aggregated proteins accumulate in the ER, certain signaling pathways such as the unfolded protein response (UPR) and ER-overload response (EOR) are functionally active in maintaining cell homeostasis. Recently we prepared Chinese hamster ovary (CHO) cells expressing mutant antithrombin (AT)(C95R) under control of the Tet-On system and showed that AT(C95R) accumulated in Russell bodies (RB), large distinctive structures derived from the ER. To characterize whether ER stress takes place in CHO cells, we examined characteristic UPR and EOR in ER stress responses. We found that the induction of ER chaperones such as Grp97, Grp78 and protein disulfide isomerase (PDI) was limited to a maximum of approximately two-fold. The processing of X-box-binding protein-1 (XBP1) mRNA and the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) subunit were not induced. Furthermore, the activation of nuclear factor-kappa B (NF-κB) was not observed. In contrast, CHO cells displayed UPR and EOR when the cells were treated with thapsigargin and tumor necrosis factor (TNF)-α, respectively. In addition, a portion of the mutant AT(C95R) was degraded through proteasomes and autophagy. CHO cells do respond to ER stress but the folding state of mutant AT(C95R) does not appear to activate the ER stress signal pathway.

Mechanisms underlying physiological functions of food factors via non-specific interactions with biological proteins

We previously reported that zerumbone, a sesquiterpene found in Zingiber zerumbet SMITH, showed notable cancer preventive effects in various organs of experimental rodents. This agent up-regulated nuclear factor-E2-related factor (Nrf2)-dependent expressions of anti-oxidative and xenobiotics-metabolizing enzymes, leading to an increased self-defense capacity. On the other hand, zerumbone markedly suppressed the expression of cyclooxygenase-2, an inducible pro-inflammatory enzyme, by disrupting mRNA stabilizing processes. Binding experiments using a biotin derivative of zerumbone demonstrated that Keap1, an Nrf2 repressive protein, is one of its major binding proteins that promotes their dissociation for inducing Nrf2 transactivation. We then generated a specific antibody against zerumbone-modified proteins and found that zerumbone modified numerous cellular proteins in a non-specific manner, with global distribution of the modified proteins seen not only in cytoplasm but also the nucleus. Based on those observations, zerumbone was speculated to cause proteo-stress, a notion supported by previous findings that it increased the C-terminus of Hsc70 interacting protein-dependent protein ubiquitination and also promoted aggresome formation. Interestingly, zerumbone counteracted proteo-stress and heat stress via up-regulation of the protein quality control systems (PQCs), e.g., heat shock proteins (HSPs), ubiquitin-proteasome, and autophagy. Meanwhile, several phytochemicals, including ursolic acid and curcumin, were identified as marked HSP70 inducers, whereas most nutrients tested were scarcely active. Recent studies have revealed that PQCs play important roles in the prevention of many lifestyle related diseases, such as cancer, thus non-specific binding of phytochemicals to cellular proteins may be a novel and unique mechanism underlying their physiological activities.

Levels of metacaspase1 and chaperones related to protein quality control in alcoholic and nonalcoholic steatohepatitis

Efficient management of misfolded or aggregated proteins in ASH and NASH is crucial for continued hepatic viability. Cellular protein quality control systems play an important role in the pathogenesis and progression of ASH and NASH. In a recent study, elevated Mca1 expression counteracted aggregation and accumulation of misfolded proteins and extended the life span of the yeast Saccharomyces cerevisiae (Hill et al, 2014). Mca1 may also associate with Ssa1 and Hsp104 in disaggregation and fragmentation of aggregated proteins and their subsequent degradation through the ER-associated degradation (ERAD) pathway. If degradation is not available, protection of the cellular environment from a misfolded protein is accomplished by its sequestration into two distinct inclusion bodies (Kaganovich et al., 2008) called the JUNQ (JUxta Nuclear Quality control compartment) and the IPOD (Insoluble Protein Deposit). Mca1, Hsp104, Hsp40, Ydj1, Ssa1, VCP/p97, and p62 all play important roles in protein quality control systems. This study aims to measure the expression of Mca1 and related chaperones involved in protein quality control in alcoholic steatohepatitis (ASH), and nonalcoholic steatohepatitis (NASH) compared with normal control liver biopsies. Mca1, Hsp104, Hsp40, Ydj1, Ssa1, VCP/p97, and p62 expressions were measured in three to six formalin-fixed paraffin embedded ASH and NASH liver biopsies and control normal liver specimens by immunofluorescence staining and quantified by immunofluorescence intensity. Mca1, Hsp104, Ydj1 and p62 were significantly upregulated compared to control (p<0.05) in ASH specimens. Hsp40 and VCP/p97 were also uptrending in ASH. In NASH, the only significant difference was the increased expression of Hsp104 compared to control (p<0.05). Ssa1 levels were uptrending in both ASH and NASH specimens. The upregulation of Mca1, Hsp104, Ydj1 and p62 in ASH may be elicited as a response to the chronic exposure of the hepatocytes to the toxicity of alcohol. Recruitment of Mca1, Hsp104, Ydj1 and p62 may indicate that autophagy, the ERAD, JUNQ, and IPOD systems are active in ASH. Whereas in NASH, elevated Hsp104 and uptrending Ssa1 levels may indicate that autophagy and IPOD may be the only active protein quality control systems involved.

Structural insight into substrate recognition by the endoplasmic reticulum folding-sensor enzyme: crystal structure of third thioredoxin-like domain of UDP-glucose:glycoprotein glucosyltransferase.

The endoplasmic reticulum (ER) possesses a protein quality control system that supports the efficient folding of newly synthesized glycoproteins. In this system, a series of N-linked glycan intermediates displayed on proteins serve as quality tags. The ER folding-sensor enzyme UDP-glucose:glycoprotein glucosyltransferase (UGGT) operates as the gatekeeper for ER quality control by specifically transferring monoglucose residues to incompletely folded glycoproteins, thereby allowing them to interact with lectin chaperone complexes to facilitate their folding. Despite its functional importance, no structural information is available for this key enzyme to date. To elucidate the folding-sensor mechanism in the ER, we performed a structural study of UGGT. Based on bioinformatics analyses, the folding-sensor region of UGGT was predicted to harbour three tandem thioredoxin (Trx)-like domains, which are often found in proteins involved in ER quality control. Furthermore, we determined the three-dimensional structure of the third Trx-like domain, which exhibits an extensive hydrophobic patch concealed by its flexible C-terminal helix. Our structural data suggest that this hydrophobic patch is involved in intermolecular interactions, thereby contributing to the folding-sensor mechanism of UGGT.

Rsp5/Nedd4 clears cells of heat-damaged proteins.

Protein quality control systems protect cells from proteotoxicity caused by the accumulation of aberrantly folded polypeptides. The Rsp5 ubiquitin ligase (mammalian homologue Nedd4) is now identified as a major constituent of a clearance pathway that degrades misfolded cytosolic proteins after exposure to heat.

Yeast prions help identify and define chaperone interaction networks.

Proteins in the cell experience various stressful conditions that can affect their ability to attain and maintain the structural conformations they need to perform effectively. Protein chaperones are an important part of a cellular protein quality control system that protects the integrity of the proteome in the face of such challenges. Chaperones from different conserved families have multiple members that cooperate to regulate each other's activity and produce machines that perform a variety of tasks. The large numbers of related chaperones with both functionally overlapping and distinct activities allows fine-tuning of the machinery for specific tasks, but presents a daunting degree of complexity. Yeast prions are misfolded forms of cellular proteins whose propagation depends on the action of protein chaperones. Studying how propagation of yeast prions is affected by alterations in functions of various chaperones provides an approach to understanding this complexity.

RNA-processing protein TDP-43 regulates FOXO-dependent protein quality control in stress response.

Protein homeostasis is critical for cell survival and functions during stress and is regulated at both RNA and protein levels. However, how the cell integrates RNA-processing programs with post-translational protein quality control systems is unknown. Transactive response DNA-binding protein (TARDBP/TDP-43) is an RNA-processing protein that is involved in the pathogenesis of major neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Here, we report a conserved role for TDP-43, from C. elegans to mammals, in the regulation of protein clearance via activation of FOXO transcription factors. In response to proteotoxic insults, TDP-43 redistributes from the nucleus to the cytoplasm, promoting nuclear translocation of FOXOs and relieving an inhibition of FOXO activity in the nucleus. The interaction between TDP-43 and the FOXO pathway in mammalian cells is mediated by their competitive binding to 14-3-3 proteins. Consistent with FOXO-dependent protein quality control, TDP-43 regulates the levels of misfolded proteins. Therefore, TDP-43 mediates stress responses and couples the regulation of RNA metabolism and protein quality control in a FOXO-dependent manner. The results suggest that compromising the function of TDP-43 in regulating protein homeostasis may contribute to the pathogenesis of related neurodegenerative diseases.

M. paratuberculosis and Parkinson’s disease – Is this a trigger

Genetic linkage studies and genome wide analysis have provided insights into complex medical diseases. Mycobacterium avium ss. paratuberculosis (MAP) causes Johne’s disease, an important enteric inflammatory disease mostly studied in ruminant animals. MAP is also the putative cause of Crohn’s disease. Moreover, MAP has been linked to other inflammatory diseases: sarcoidosis, Blau syndrome, autoimmune diabetes, autoimmune thyroiditis and multiple sclerosis. Genetic studies reveal an association between Parkinson’s disease (PD), leprosy and Crohn’s disease and since discovered, these findings have been considered “surprising”. Autophagy and ubiquitin–proteosome systems are cellular systems that both fight intracellular pathogens (xenophagy) and maintain cellular protein quality control. PD is a common neurodegenerative disease that manifests clinically as a profound movement disorder. The recognized genetic defects of PD create disruption of cellular homeostasis that result in protein folding abnormalities of PD called Lewy bodies. Those same genetic defects are associated with susceptibility to intracellular pathogens, including mycobacteria. It is now understood that PD Lewy body pathology starts in the enteric nervous system and “spreads” to the brain in a retrograde fashion via the vagus nerve. This is the same process by which prions affect the brain. Lewy body pathology of the enteric nervous system predates the Lewy body pathology of the central nervous system (CNS) by years or even decades. This article proposes that genetic defects associated with PD also result in a permissive environment for MAP infection – ineffective xenophagy. It postulates that beginning as an enteric infection, MAP – via the vagus nerve – initiates a pathologic process that results in a targeted neuroinvasion of the CNS. The article proposes that MAP infection and resultant PD pathology are due, in the genetically at-risk and age dependant, to the consumptive exhaustion of the protein quality control systems.

Muscle proteomics reveals novel insights into the pathophysiological mechanisms of collagen VI myopathies.

Mutations in the collagen VI genes cause the Ullrich congenital muscular dystrophy (UCMD), with severe phenotype, and Bethlem myopathy (BM) with mild to moderate phenotype. Both, UCMD and BM patients show dystrophic features with degeneration/regeneration and replacement of muscle with fat and fibrous connective tissue. At molecular level, UCMD patients show autophagic impairment and increased PTP opening; these features are less severe in BM. To elucidate the biochemical mechanisms adopted by the muscle to adapt to collagen VI deficiency in BM and UCMD patients, a proteome analysis was carried out on human muscle biopsies. Qualitative and quantitative differences were assessed by 2D-DIGE coupled to MALDI-ToF/ToF MS. Proteomics results, coupled with immunoblotting, indicate changes in UPR, hexosamine pathway, and amino acid and fatty acid metabolism, suggesting an association of ER stress, metabolic dysregulation, autophagic impairment, and alteration in mechanotransduction signaling. Overall, these results indicate that despite the common downregulation of hexosamine pathway in UCMD and BM, in BM the protein quality control system is sustained by a metabolic adaptation supporting energy requirements for the maintenance of autophagy, counteracting ER misfolded protein overload. In UCMD, this multilayered system may be disrupted and worsened by the metabolic rewiring, which leads to lipotoxicity.

Profiling the mitochondrial proteome of Leber's Hereditary Optic Neuropathy (LHON) in Thailand: down-regulation of bioenergetics and mitochondrial protein quality control pathways in fibroblasts with the 11778G>A mutation.

Leber’s Hereditary Optic Neuropathy (LHON) is one of the commonest mitochondrial diseases. It causes total blindness, and predominantly affects young males. For the disease to develop, it is necessary for an individual to carry one of the primary mtDNA mutations 11778G>A, 14484T>C or 3460G>A. However these mutations are not sufficient to cause disease, and they do not explain the characteristic features of LHON such as the higher prevalence in males, incomplete penetrance, and relatively later age of onset. In order to explore the roles of nuclear encoded mitochondrial proteins in development of LHON, we applied a proteomic approach to samples from affected and unaffected individuals from 3 pedigrees and from 5 unrelated controls. Two-dimensional electrophoresis followed by MS/MS analysis in the mitochondrial lysate identified 17 proteins which were differentially expressed between LHON cases and unrelated controls, and 24 proteins which were differentially expressed between unaffected relatives and unrelated controls. The proteomic data were successfully validated by western blot analysis of 3 selected proteins. All of the proteins identified in the study were mitochondrial proteins and most of them were down regulated in 11778G>A mutant fibroblasts. These proteins included: subunits of OXPHOS enzyme complexes, proteins involved in intermediary metabolic processes, nucleoid related proteins, chaperones, cristae remodelling proteins and an anti-oxidant enzyme. The protein profiles of both the affected and unaffected 11778G>A carriers shared many features which differed from those of unrelated control group, revealing similar proteomic responses to 11778G>A mutation in both affected and unaffected individuals. Differentially expressed proteins revealed two broad groups: a cluster of bioenergetic pathway proteins and a cluster involved in protein quality control system. Defects in these systems are likely to impede the function of retinal ganglion cells, and may lead to the development of LHON in synergy with the primary mtDNA mutation.

Structural features of free N-glycans occurring in plants and functional features of de-N-glycosylation enzymes, ENGase, and PNGase: the presence of unusual plant complex type N-glycans.

Free N-glycans (FNGs) are present at micromolar concentrations in plant cells during their differentiation, growth, and maturation stages. It has been postulated that these FNGs are signaling molecules involved in plant development or fruit ripening. However, the hypothetical biochemical and molecular function of FNGs has not been yet established. The structure of FNGs found ubiquitously in plant tissues such as hypocotyls, leaves, roots, developing seeds, or fruits can be classified into two types: high-mannose type and plant complex type; the former, in most cases, has only one GlcNAc residue at the reducing end (GN1 type), while the latter has the chitobiosyl unit at the reducing end (GN2 type). These findings suggest that endo-β-N-acetylglucosaminidase (ENGase) must be involved in the production of GN1 type FNGs, whereas only peptide:N-glycanase (PNGase) is involved in the production of GN2 type FNGs. It has been hypothesized that cytosolic PNGase (cPNGase) and ENGase in animal cells are involved in the production of high-mannose type FNGs in order to release N-glycans from the misfolded glycoproteins in the protein quality control systems. In the case of plants, it is well known that another type of PNGase, the acidic PNGase (aPNGase) is involved in the production of plant complex type FNGs in an acidic organelle, suggesting the de-N-glycosylation mechanism in plants is different from that in animal cells. To better understand the role of these FNGs in plants, the genes encoding these N-glycan releasing enzymes (ENGase and PNGase) were first identified, and then structure of FNGs in ENGase knocked-out plants were analyzed. These transgenic plants provide new insight into the plant-specific de-N-glycosylation mechanism and putative physiological functions of FNGs. In this review, we focus on the structural features of plant FNGs, as well as functional features of cPNGase/ENGase and plant specific PNGase, and putative functions of FNGs are also discussed.

A.I.4: Protein aggregate myopathies: The many faces of an expanding disease group

Pathological protein aggregates are the characteristic feature in a wide variety of human diseases affecting the central and peripheral nervous system, eyes, skin, liver as well as heart and skeletal muscles. Protein aggregate myopathies (PAM) are an emerging and numerically significant group of hereditary and acquired striated muscle diseases with marked clinical and genetic heterogeneity. Their disease manifestation varies from the neonatal period to late adulthood. They are present in diverse ethnic groups, affect both sexes, and their clinical presentation ranges from congenital myopathies over scapuloperoneal and oculopharyngeal phenotypes to distal, limb girdle, and generalized patterns of muscle weakness and wasting. PAM are due to mutations in genes coding for cytoskeletal proteins, which are either components of the myofibrillar apparatus (α-actin, myosins, titin, filamin C, myotilin, ZASP, BAG-3, FHL1) or the three-dimensional filamentous extrasarcomeric network (desmin, plectin). In addition, mutations in genes coding for proteins with essential functions in protein quality control (VCP, DNAJB6, BAG-3, aB-crystallin), selenoproteins (selenoprotein N), and DNA-binding proteins (PABPN1) have been described. Exciting novel insight into the molecular pathogenesis of PAM lead to concepts in which PAM-related gene defects cause effects at multiple levels by influencing the structure and function of motor proteins, sarcomeric and extrasarcomeric cytoskeleton proteins, signaling cascades, protein quality control systems as well as the function of cell organelles in striated muscle cells. The presentation will give an overview on the clinical, myopathological and pathophysiological aspects of PAM. In addition, central results from the analysis of novel cell and animal models, the implementation of a PAM patient registry and a pilot study for the treatment of PAM will be highlighted. Pathological protein aggregates are the characteristic feature in a wide variety of human diseases affecting the central and peripheral nervous system, eyes, skin, liver as well as heart and skeletal muscles. Protein aggregate myopathies (PAM) are an emerging and numerically significant group of hereditary and acquired striated muscle diseases with marked clinical and genetic heterogeneity. Their disease manifestation varies from the neonatal period to late adulthood. They are present in diverse ethnic groups, affect both sexes, and their clinical presentation ranges from congenital myopathies over scapuloperoneal and oculopharyngeal phenotypes to distal, limb girdle, and generalized patterns of muscle weakness and wasting. PAM are due to mutations in genes coding for cytoskeletal proteins, which are either components of the myofibrillar apparatus (α-actin, myosins, titin, filamin C, myotilin, ZASP, BAG-3, FHL1) or the three-dimensional filamentous extrasarcomeric network (desmin, plectin). In addition, mutations in genes coding for proteins with essential functions in protein quality control (VCP, DNAJB6, BAG-3, aB-crystallin), selenoproteins (selenoprotein N), and DNA-binding proteins (PABPN1) have been described. Exciting novel insight into the molecular pathogenesis of PAM lead to concepts in which PAM-related gene defects cause effects at multiple levels by influencing the structure and function of motor proteins, sarcomeric and extrasarcomeric cytoskeleton proteins, signaling cascades, protein quality control systems as well as the function of cell organelles in striated muscle cells. The presentation will give an overview on the clinical, myopathological and pathophysiological aspects of PAM. In addition, central results from the analysis of novel cell and animal models, the implementation of a PAM patient registry and a pilot study for the treatment of PAM will be highlighted.

P62/sequestosome 1 regulates aggresome formation of pathogenic ataxin-3 with expanded polyglutamine.

The cellular protein quality control system in association with aggresome formation contributes to protecting cells against aggregation-prone protein-induced toxicity. p62/Sequestosome 1 (p62) is a multifunctional protein which plays an important role in protein degradation and aggregation. Although poly-ubiquitination is usually required for p62-mediated protein degradation and aggresome formation, several p62 substrates are processed to form aggregate in an ubiquitination-independent manner. In this study we demonstrate that p62 directly interacts with pathogenic Machado Joseph Disease (MJD)-associated protein ataxin-3 with polyglutamine (polyQ) expansion. Moreover, p62 could regulate the aggresome formation of pathogenic ataxin-3 and protect cells against pathogenic ataxin-3-induced cell death.

Predicting aggregation-prone sequences in proteins.

Owing to its association with a diverse range of human diseases, the determinants of protein aggregation are studied intensively. It is generally accepted that the effective aggregation tendency of a protein depends on many factors such as folding efficiency towards the native state, thermodynamic stability of that conformation, intrinsic aggregation propensity of the polypeptide sequence and its ability to be recognized by the protein quality control system. The intrinsic aggregation propensity of a polypeptide sequence is related to the presence of short APRs (aggregation-prone regions) that self-associate to form intermolecular β-structured assemblies. These are typically short sequence segments (5-15 amino acids) that display high hydrophobicity, low net charge and a high tendency to form β-structures. As the presence of such APRs is a prerequisite for aggregation, a plethora of methods have been developed to identify APRs in amino acid sequences. In the present chapter, the methodological basis of these approaches is discussed, as well as some practical applications.

Protein folding, misfolding and quality control: the role of molecular chaperones.

Cells have to cope with stressful conditions and adapt to changing environments. Heat stress, heavy metal ions or UV stress induce damage to cellular proteins and disturb the balanced status of the proteome. The adjusted balance between folded and folding proteins, called protein homoeostasis, is required for every aspect of cellular functionality. Protective proteins called chaperones are expressed under extreme conditions in order to prevent aggregation of cellular proteins and safeguard protein quality. These chaperones co-operate during de novo folding, refolding and disaggregation of damaged proteins and in many cases refold them to their functional state. Even under physiological conditions these machines support protein homoeostasis and maintain the balance between de novo folding and degradation. Mutations generating unstable proteins, which are observed in numerous human diseases such as Alzheimer's disease, Huntington's disease, amyotrophic lateral sclerosis and cystic fibrosis, also challenge the protein quality control system. A better knowledge of how the protein homoeostasis system is regulated will lead to an improved understanding of these diseases and provide potential targets for therapy.

Oxidative Stress and Proteostasis Network: Culprit and Casualty of Alzheimer’s-Like Neurodegeneration

Free radical-mediated damage to proteins is particularly important in aging and age-related neurodegenerative diseases, because in the majority of cases it is a non-reversible phenomenon that requires clearance systems for removal. Major consequences of protein oxidation are loss of protein function and the formation of large protein aggregates, which are often toxic to cells if allowed to accumulate. Deposition of aggregated, misfolded, and oxidized proteins may also result from the impairment of protein quality control (PQC) system, including protein unfolded response, proteasome, and autophagy. Perturbations of such components of the proteostasis network that provides a critical protective role against stress conditions are emerging as relevant factor in triggering neuronal death. In this outlook paper, we discuss the role of protein oxidation as a major contributing factor for the impairment of the PQC regulating protein folding, surveillance, and degradation. Recent studies from our group and from others aim to better understand the link between Down syndrome and Alzheimer’s disease neuropathology. We propose oxidative stress and alteration of proteostasis network as a possible unifying mechanism triggering neurodegeneration.

Unstable mutant lysozymes are degraded through the interaction with calnexin homolog Cne1p in Saccharomyces cerevisiae.

Cne1p is a yeast homolog of calnexin, which is a constituent of endoplasmic reticulum (ER)-associated protein quality control system in mammals. Cne1p may be involved in the degradation of misfolded lysozymes in Saccharomyces cerevisiae. To test this, c-Myc-tagged lysozymes were expressed in CNE1-deficient S. cerevisiae. The expression and secretion of an unstable lysozyme mutant G49N/D66H were enhanced and its intracellular localization was changed in the CNE1-deficient strain. Furthermore, when Cne1p was co-expressed with unstable lysozyme mutants (G49N/D66H, G49N/C76A, and K13D/G49N), its affinity to the misfolded mutant proteins was revealed by co-immunoprecipitation. The interaction with Cne1p was abrogated by the addition of tunicamycin, an inhibitor of N-glycosylation, indicating that N-linked carbohydrates might be necessary for protein binding to Cne1p. These results suggest that in yeasts, Cne1p interacts with misfolded lysozyme proteins possibly causing their retention in the ER and subsequent elimination via ER-associated degradation.

Analysis of modifications in the ER protein quality control system on heterologous protein expression in Saccharomyces cerevisiae

Analysis of modifications in the ER protein quality control system on heterologous protein expression in Saccharomyces cerevisiae