Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P11021 (BiP)
 2,049 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The endoplasmic reticulum (ER) quality-control machinery maintains the fidelity of the maturation process by sorting aberrant proteins for ER-associated protein degradation (ERAD), a process requiring retrotranslocation from the ER lumen to the cytosol and degradation by the proteasome. Here, we assessed the role of N-linked glycans in ERAD by monitoring the degradation of wild-type (Tyr) and albino mutant (Tyr(C85S)) tyrosinase. Initially, mutant tyrosinase was established as a genuine ERAD substrate using intact melanocyte and semi-permeabilized cell systems. Inhibiting mannose trimming or accumulating Tyr(C85S) in a monoglucosylated form led to its stabilization, supporting a role for lectin chaperones in ER retention and proteasomal degradation. In contrast, ablating the lectin chaperone interactions by preventing glucose trimming caused a rapid disappearance of tyrosinase, initially due to the formation of protein aggregates, which were subsequently degraded by the proteasome. The co-localization of aggregated tyrosinase with protein disulfide isomerase and BiP, but not calnexin, supports an ER organization, which aids in protein maturation and degradation. Based on these studies, we propose a model of tyrosinase degradation in which interactions between N-linked glycans and lectin chaperones help to minimize tyrosinase aggregation and also target non-native substrates for retro-translocation and subsequent degradation.
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PMID:Carbohydrates act as sorting determinants in ER-associated degradation of tyrosinase. 1516 41

Apoptosis is probably the main form of beta-cell death in both type 1 diabetes mellitus (T1DM) and T2DM. In T1DM, cytokines contribute to beta-cell destruction through nuclear factor-kappaB (NF-kappaB) activation. Previous studies suggested that in T2DM high glucose and free fatty acids (FFAs) are beta-cell toxic also via NF-kappaB activation. The aims of this study were to clarify whether common mechanisms are involved in FFA- and cytokine-induced beta-cell apoptosis and determine whether TNFalpha, an adipocyte-derived cytokine, potentiates FFA toxicity through enhanced NF-kappaB activation. Apoptosis was induced in insulinoma (INS)-1E cells, rat islets, and fluorescence-activated cell sorting-purified beta-cells by oleate, palmitate, and/or cytokines (IL-1beta, interferon-gamma, TNFalpha). Palmitate and IL-1beta induced a similar percentage of apoptosis in INS-1E cells, whereas oleate was less toxic. TNFalpha did not potentiate FFA toxicity in primary beta-cells. The NF-kappaB-dependent genes inducible nitric oxide synthase and monocyte chemoattractant protein-1 were induced by IL-1beta but not by FFAs. Cytokines activated NF-kappaB in INS-1E and beta-cells, but FFAs did not. Moreover, FFAs did not enhance NF-kappaB activation by TNFalpha. Palmitate and oleate induced C/EBP homologous protein, activating transcription factor-4, and immunoglobulin heavy chain binding protein mRNAs, X-box binding protein-1 alternative splicing, and activation of the activating transcription factor-6 promoter in INS-1E cells, suggesting that FFAs trigger an endoplasmic reticulum (ER) stress response. We conclude that apoptosis is the main mode of FFA- and cytokine-induced beta-cell death but the mechanisms involved are different. Whereas cytokines induce NF-kappaB activation and ER stress (secondary to nitric oxide formation), FFAs activate an ER stress response via an NF-kappaB- and nitric oxide-independent mechanism. Our results argue against a unifying hypothesis for the mechanisms of beta-cell death in T1DM and T2DM.
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PMID:Free fatty acids and cytokines induce pancreatic beta-cell apoptosis by different mechanisms: role of nuclear factor-kappaB and endoplasmic reticulum stress. 1529 38

Metabolic labeling studies were conducted in freshly isolated mouse islets and a beta-cell line (MIN6) to examine the effects of proteasome inhibition on glucose-stimulated (pro)insulin synthesis and secretion. Glucose-stimulated (pro)insulin synthesis, as determined by the incorporation of [(3)H]tyrosine, decreased significantly by 90% in islets and 71% in MIN6 cells pretreated with the proteasome inhibitor lactacystin (10 microM) for 2 h. To follow the fate of newly synthesized (pro)insulin, islets were pulse-labeled with [(3)H]tyrosine (40 microCi) for 20 min and chased +/- lactacystin (10 microM) for up to 4 h. The release of newly synthesized (pro)insulin ([(3)H]tyrosine-labeled) was similar between lactacystin-treated and control islets despite a 51% decrease (p <0.05) in total immunoreactive (pro)insulin secretion by lactacystin-treated islets. The specific radioactivity of [(3)H]tyrosine-labeled (pro)insulin in the extracellular medium of lactacystin-treated islets (0.52 +/- 0.16 cpm/microunits) was 2-fold greater relative to control islets (0.25 +/- 0.06 cpm/microunits). Induction of the unfolded protein response by lactacystin, as evidenced by the up-regulation of endoplasmic reticulum (ER) chaperones (GRP78/BiP, GRP94, protein disulfide isomerase) and induction of the stress-inducible transcription factor C/EBP-homologous protein/GADD153 (CHOP/GADD153), likely contributed to the release of newly synthesized (pro)insulin to relieve ER stress. The present data indicate proteasome inhibition did not prevent, but increased (p <0.05), the intracellular degradation of [(3)H]tyrosine-labeled (pro-)insulin from 8 to 24% in islets. Collectively, these data indicate beta-cells may balance glucose-stimulated (pro)insulin synthesis and secretion with the activity of the proteasome to regulate protein concentrations in the ER.
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PMID:Proteasome inhibition alters glucose-stimulated (pro)insulin secretion and turnover in pancreatic {beta}-cells. 1570 91

The mechanism of how fluoride causes fluorosis remains unknown. Exposure to fluoride can inhibit protein synthesis, and this may also occur by agents that cause endoplasmic reticulum (ER) stress. When translated proteins fail to fold properly or become misfolded, ER stress response genes are induced that together comprise the unfolded protein response. Because ameloblasts are responsible for dental enamel formation, we used an ameloblast-derived cell line (LS8) to characterize specific responses to fluoride treatment. LS8 cells were growth-inhibited by as little as 1.9-3.8 ppm fluoride, whereas higher doses induced ER stress and caspase-mediated DNA fragmentation. Growth arrest and DNA damage-inducible proteins (GADD153/CHOP, GADD45alpha), binding protein (BiP/glucose-responsive protein 78 (GRP78), the non-secreted form of carbonic anhydrase VI (CA-VI), and active X-box-binding protein-1 (Xbp-1) were all induced significantly after exposure to 38 ppm fluoride. Unexpectedly, DNA fragmentation increased when GADD153 expression was inhibited by short interfering RNA treatment but remained unaffected by transient GADD153 overexpression. Analysis of control and GADD153(-/-) embryonic fibroblasts demonstrated that caspase-3 mediated the increased DNA fragmentation observed in the GADD153 null cells. We also demonstrate that mouse incisor ameloblasts are sensitive to the toxic effects of high dose fluoride in drinking water. Activated Ire1 initiates an ER stress response pathway, and mouse ameloblasts were shown to express activated Ire1. Ire1 levels appeared induced by fluoride treatment, indicating that ER stress may play a role in dental fluorosis. Low dose fluoride, such as that present in fluoridated drinking water, did not induce ER stress.
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PMID:Fluoride induces endoplasmic reticulum stress in ameloblasts responsible for dental enamel formation. 1584 62

Numerous xenobiotics, including therapeutics agents, are substrates for bioactivation to electrophilic reactive intermediates that may covalently modify biomolecules. Selective estrogen receptor modulators (SERMs) are in clinical use for long-term therapy of postmenopausal syndromes and chemoprevention and provide a potential alternative for hormone replacement therapy (HRT). Raloxifene, in common with many SERMs and other xenobiotics, is a polyaromatic phenol that has been shown to be metabolically bioactivated to electrophilic and redox active quinoids. Nucleic acid and glutathione adduct formation have been reported, but little is known about protein covalent modification. A novel COATag (covert oxidatively activated tag) was synthesized in which raloxifene was linked to biotin. The COATag was reactive toward a model protein, human glutathione-S-transferase P1-1, in the presence but not the absence of monooxygenase. The covalent modification of proteins in rat liver microsomal incubations was NADPH-dependent implicating cytochrome P450 oxidase. The COATag facilitated isolation and identification of covalently modified microsomal proteins: cytosolic glucose regulated protein (GRP78/BiP), three protein disulfide isomerases, and microsomal glutathione S-transferase 1. Oxidative metabolism of raloxifene produces reactive intermediates of sufficient lifetimes to covalently modify proteins in tissue microsomes, behavior anticipated for other polyaromatic phenol xenobiotics that can be tested by the COATag methodology. The combined use of a COATag with a simple biotin-linked electrophile (such as an iodoacetamide tag) is a new technique that allows quantification of protein covalent modification via alkylation vs oxidation in response to xenobiotic reactive intermediates. The identification of modified proteins is important for defining pathways that might lead alternatively to either cytotoxicity or cytoprotection.
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PMID:Analysis of protein covalent modification by xenobiotics using a covert oxidatively activated tag: raloxifene proof-of-principle study. 1616 42

Recently, it has become apparent that asparagine-linked (N-linked) oligosaccharide at an early stage of processing can play an important role in quality control of the secretory pathway. Here, we have developed a system for better understanding of the N-glycosylation machinery and its involvement in quality control in the endoplasmic reticulum (ER). Rough microsomes (RM) treated with 0.18% Tx-100 (TxRM) preserved translocation activities to a similar extent detected in RM. TxRM were depleted of many soluble proteins including glucosidase II, BiP and Erp72, but maintained approximately 80% of calnexin, a membrane protein. More importantly, TxRM revealed insufficient glycosylation of T cell receptor-alpha (TCR-alpha), suggesting that a factor or factors extracted with 0.18% Tx-100 is responsible for facilitating the transfer of oligosaccharides to the protein. In addition, the top band of TCR-alpha translated in TxRM migrated slower than that in RM, but faster than that in RM treated with castanospermine (CST), an inhibitor of glucosidase I/II. This suggests that the trimming of the inner two glucose sugars is impaired by the loss of glucosidase II. Furthermore, we demonstrated that TCR-alpha coprecipitated with calnexin migrated between unglucosylated and diglucosylated forms on SDS-PAGE. Thus, the treatment of RM with low concentration of detergent is a very powerful method for elucidating not only N-glycosylation processes but also other biological functions such as quality control in the ER.
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PMID:A novel approach for N-glycosylation studies using detergent extracted microsomes. 1618 Jan 1

The folding of glycoproteins in the endoplasmic reticulum (ER) depends on a quality control mechanism mediated by the calnexin/calreticulin cycle. During this process, continuous glucose trimming and UDP-glucose-dependent re-glucosylation of unfolded glycoproteins takes place. To ensure proper folding, increases in misfolded proteins lead to up-regulation of the components involved in quality control through a process known as the unfolded protein response (UPR). Reglucosylation is catalyzed by the ER lumenal located enzyme UDP-glucose glycoprotein glucosyltransferase, but as UDP-glucose is synthesized in the cytosol, a UDP-glucose transporter is required in the calnexin/calreticulin cycle. Even though such a transporter has been hypothesized, no protein playing this role in the ER yet has been identified. Here we provide evidence that AtUTr1, a UDP-galactose/glucose transporter from Arabidopsis thaliana, responds to stimuli that trigger the UPR increasing its expression around 9-fold. The accumulation of AtUTr1 transcript is accompanied by an increase in the level of the AtUTr1 protein. Moreover, subcellular localization studies indicate that AtUTr1 is localized in the ER of plant cells. We reasoned that an impairment in AtUTr1 expression should perturb the calnexin/calreticulin cycle leading to an increase in misfolded protein and triggering the UPR. Toward that end, we analyzed an AtUTr1 insertional mutant and found an up-regulation of the ER chaperones BiP and calnexin, suggesting that these plants may be constitutively activating the UPR. Thus, we propose that in A. thaliana, AtUTr1 is the UDP-glucose transporter involved in quality control in the ER.
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PMID:AtUTr1, a UDP-glucose/UDP-galactose transporter from Arabidopsis thaliana, is located in the endoplasmic reticulum and up-regulated by the unfolded protein response. 1646 98

BiP is a molecular chaperone induced in the unfolded protein response (UPR). In mammalian cells, BiP is induced by glucose starvation when it is called glucose-regulated protein 78 (GRP78). In Arabidopsis thaliana, however, we demonstrated that BiP transcripts decreased with sugar depletion and increased with sugar addition. Transcripts for beta-glucuronidase (GUS) driven by BiP promoter respond to tunicamycin and sugar, being similar with endogenous BiP transcripts in transgenic A. thaliana. When GUS was regulated by P-UPRE, a cis-element responsible for the UPR identified in BiP promoter, GUS transcripts were accumulated by sugar starvation. Subsequently, transgenic A. thaliana harboring luciferase (LUC) gene regulated by P-UPRE was analyzed. Sugar depletion also increased LUC activity. It is concluded that BiP is induced by sugar independent of the cis-element responsible for the UPR.
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PMID:Induction of BiP by sugar independent of a cis-element for the unfolded protein response in Arabidopsis thaliana. 1678 68

In pancreatic beta cells, the endoplasmic reticulum (ER) is an important site for insulin biosynthesis and the folding of newly synthesized proinsulin. Here, we show that IRE1alpha, an ER-resident protein kinase, has a crucial function in insulin biosynthesis. IRE1alpha phosphorylation is coupled to insulin biosynthesis in response to transient exposure to high glucose; inactivation of IRE1alpha signaling by siRNA or inhibition of IRE1alpha phosphorylation hinders insulin biosynthesis. IRE1 activation by high glucose does not accompany XBP-1 splicing and BiP dissociation but upregulates its target genes such as WFS1. Thus, IRE1 signaling activated by transient exposure to high glucose uses a unique subset of downstream components and has a beneficial effect on pancreatic beta cells. In contrast, chronic exposure of beta cells to high glucose causes ER stress and hyperactivation of IRE1, leading to the suppression of insulin gene expression. IRE1 signaling is therefore a potential target for therapeutic regulation of insulin biosynthesis.
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PMID:Regulation of insulin biosynthesis in pancreatic beta cells by an endoplasmic reticulum-resident protein kinase IRE1. 1695 Jan 32

Mammalian transcription factor ATF6 is constitutively synthesized as a type II transmembrane protein embedded in the endoplasmic reticulum (ER). Upon ER stress ATF6 is transported to the Golgi apparatus where it is cleaved to release its cytoplasmic domain. This is then translocated into the nucleus where it activates transcription of ER-localized molecular chaperones and folding enzymes to maintain the homeostasis of the ER. We recently found that, owing to the presence of intra- and intermolecular disulfide bridges, ATF6 occurs in unstressed ER in monomer, dimer and oligomer forms. Disulfide-bonded ATF6 is reduced on treatment of cells with various chemical ER stress inducers, and only the reduced monomer ATF6 reaches the Golgi apparatus. In this study, we evoked ER stress under more physiological conditions, namely, glucose starvation, and analyzed its consequence for ATF6 activation. Glucose starvation activated ATF6 and induced the ER chaperone BiP, albeit weakly. ATF6 was thus dissociated from BiP, transported to the Golgi apparatus, and cleaved. Glucose starvation enhanced the synthesis of ATF6 approximately two-fold, probably via transcriptional induction. Importantly, reduction of disulfide bridges and transport of reduced monomer occurred in response to glucose starvation. We conclude that ER stress-induced reduction of ATF6 represents a general feature of the ATF6 activation process.
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PMID:Reduction of disulfide bridges in the lumenal domain of ATF6 in response to glucose starvation. 1790 5


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