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)

Protein folding and quality control in the early secretory pathway function as posttranslational checkpoints in eukaryote gene expression. Herein, an aberrant form of the hepatic secretory protein alpha1-antitrypsin was stably expressed in a human embryonic kidney cell line to elucidate the mechanisms by which glycoprotein endoplasmic reticulum-associated degradation (GERAD) is administered in cells from higher eukaryotes. After biosynthesis, genetic variant PI Z underwent alternative phases of secretion and degradation, the latter of which was mediated by the proteasome. Degradation required release from calnexin- and asparagine-linked oligosaccharide modification by endoplasmic reticulum mannosidase I, the latter of which occurred as PI Z was bound to the molecular chaperone grp78/BiP. That a distinct GERAD program operates in human embryonic kidney cells was supported by the extent of PI Z secretion, apparent lack of polymerization, inability of calnexin to participate in the degradation process, and sequestration of the glycoprotein folding sensor UDP-glucose:glycoprotein glucosyltransferase in the Golgi complex. Because UDP-glucose:glycoprotein glucosyltransferase sustains calnexin binding, its altered distribution is consistent with a GERAD program that hinders the reentry of substrates into the calnexin cycle, allowing grp78/BiP to partner with a lectin, other than calnexin, in the recognition of a two-component GERAD signal to facilitate substrate recruitment. How the processing of a mutant protein, rather than the mutation itself, can contribute to disease pathogenesis, is discussed.
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PMID:Organizational diversity among distinct glycoprotein endoplasmic reticulum-associated degradation programs. 1218 35

The thyrotropin receptor (TSHR) is a member of the G protein-coupled receptor superfamily. It has by now been clearly established that the maturation of the glycoproteins synthesized in the endoplasmic reticulum involves interactions with molecular chaperones, which promote the folding and assembly of the glycoproteins. In this study, we investigated whether calnexin (CNX), calreticulin (CRT) and BiP, three of the main molecular chaperones present in the endoplasmic reticulum, interact with the TSHR and what effects these interactions might have on the folding of the receptor. In the first set of experiments, we observed that in a K562 cell line expressing TSHR, about 50% of the receptor synthesized was degraded by the proteasome after ubiquitination. In order to determine whether TSHR interact with CNX, CRT and BiP, coimmunoprecipitation experiments were performed. TSHR was found to be associated with all three molecular chaperones. To study the role of the interactions between CNX and CRT and the TSHR, we used castanospermine, a glucosidase I and II inhibitor that blocks the interactions between these chaperones and glycoproteins. In K562 cells expressing the TSHR, these drugs led to a faster degradation of the receptor, which indicates that these interactions contribute to stabilizing the receptor after its synthesis. The overexpression of calnexin and calreticulin in these cells stabilizes the receptor during the first hour after its synthesis, whereas the degradation of TSHR increased in a cell line overexpressing BiP and the quantity of TSHR able to acquire complex type oligosaccharides decreased. These results show that calnexin, calreticulin and BiP all interact with TSHR and that the choice made between these two chaperone systems is crucial because each of them has distinct effects on the folding and stability of this receptor at the endoplasmic reticulum level.
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PMID:Association of the thyrotropin receptor with calnexin, calreticulin and BiP. Efects on the maturation of the receptor. 1238 51

During its initial folding in the endoplasmic reticulum (ER), newly synthesized thyroglobulin (Tg) is known to interact with calnexin and other ER molecular chaperones, but its interaction with calreticulin has not been examined previously. In the present study, we have investigated the interactions of endogenous Tg with calreticulin and with several other ER chaperones. We find that, in FRTL-5 and PC-Cl3 cells, calnexin and calreticulin interact with newly synthesized Tg in a carbohydrate-dependent manner, with largely overlapping kinetics that are concomitant with the maturation of Tg intrachain disulphide bonds, preceding Tg dimerization and exit from the ER. Calreticulin co-precipitates more newly synthesized Tg than does calnexin; however, using two different experimental approaches, calnexin and calreticulin were found in ternary complexes with Tg, making this the first endogenous protein reported in ternary complexes with calnexin and calreticulin in the ER of live cells. Depletion of Ca(2+) from the ER elicited by thapsigargin (a specific inhibitor of ER Ca(2+)-ATPases) results in retention of Tg in this organelle. Interestingly, thapsigargin treatment induces the premature exit of Tg from the calnexin/calreticulin cycle, while stabilizing and prolonging interactions of Tg with BiP (immunoglobulin heavy chain binding protein) and GRP94 (glucose-regulated protein 94), two chaperones whose binding is not carbohydrate-dependent. Our results suggest that calnexin and calreticulin, acting in ternary complexes with a large glycoprotein substrate such as Tg, might be engaged in the folding of distinct domains, and indicate that lumenal Ca(2+) strongly influences the folding of exportable glycoproteins, in part by regulating the balance of substrate binding to different molecular chaperone systems within the ER.
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PMID:Folding of thyroglobulin in the calnexin/calreticulin pathway and its alteration by loss of Ca2+ from the endoplasmic reticulum. 1240 Nov 14

We demonstrate the existence of a large endoplasmic reticulum (ER)-localized multiprotein complex that is comprised of the molecular chaperones BiP; GRP94; CaBP1; protein disulfide isomerase (PDI); ERdj3, a recently identified ER Hsp40 cochaperone; cyclophilin B; ERp72; GRP170; UDP-glucosyltransferase; and SDF2-L1. This complex is associated with unassembled, incompletely folded immunoglobulin heavy chains. Except for ERdj3, and to a lesser extent PDI, this complex also forms in the absence of nascent protein synthesis and is found in a variety of cell types. Cross-linking studies reveal that the majority of these chaperones are included in the complex. Our data suggest that this subset of ER chaperones forms an ER network that can bind to unfolded protein substrates instead of existing as free pools that assembled onto substrate proteins. It is noticeable that most of the components of the calnexin/calreticulin system, which include some of the most abundant chaperones inside the ER, are either not detected in this complex or only very poorly represented. This study demonstrates an organization of ER chaperones and folding enzymes that has not been previously appreciated and suggests a spatial separation of the two chaperone systems that may account for the temporal interactions observed in other studies.
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PMID:A subset of chaperones and folding enzymes form multiprotein complexes in endoplasmic reticulum to bind nascent proteins. 1247 65

Apolipoprotein[a] (apo[a]) is a large disulfide linked glycoprotein synthesized by hepatocytes. We have examined the role of disulfide bond formation in the processing of apo[a] using human and rat hepatoma cells expressing apo[a] isoforms containing varying numbers of kringle 4 (K4) domains, following treatment with DTT. Hepatoma cells expressing 6- or 9-K4 isoforms revealed approximately 90% inhibition of apo[a] secretion following DTT treatment, although larger isoforms containing 13- or 17-K4 domains demonstrated continued secretion (up to 30% of control values), suggesting that a fraction of the larger isoforms is at least partially DTT resistant. Wash-out experiments demonstrated that these effects were completely reversible for all isoforms studied, with no enhanced degradation associated with prolonged intracellular retention. DTT treatment was associated with enhanced binding of apo[a] with the endoplasmic reticulum-associated chaperone proteins calnexin, calreticulin, and BiP, which was reversible upon DTT removal. The chemical chaperone 6-aminohexanoic acid, previously demonstrated by others to rescue defective apo[a] secretion associated with alterations in glycosylation, failed to alter the secretion of apo[a] following DTT treatment. The demonstration that DTT modulates apo[a] secretion in a manner influenced by both the type and number of K4 repeats extends understanding of the mechanisms that regulate its exit from the endoplasmic reticulum.
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PMID:Apolipoprotein[a] secretion from hepatoma cells is regulated in a size-dependent manner by alterations in disulfide bond formation. 1256 43

Dendritic cells (DCs) are likely to play a key role in immunity against Mycobacterium tuberculosis, but the fate of the bacterium in these cells is still unknown. Here we report that, unlike macrophages (Mphis), human monocyte-derived DCs are not permissive for the growth of virulent M. tuberculosis H37Rv. Mycobacterial vacuoles are neither acidic nor fused with host cell lysosomes in DCs, in a mode similar to that seen in mycobacterial infection of Mphis. However, uptake of the fluid phase marker dextran, and of transferrin, as well as accumulation of the recycling endosome-specific small GTPase Rab11 onto the mycobacterial phagosome, are almost abolished in infected DCs, but not in Mphis. Moreover, communication between mycobacterial phagosomes and the host-cell biosynthetic pathway is impaired, given that <10% of M. tuberculosis vacuoles in DCs stained for the endoplasmic reticulum-specific proteins Grp78/BiP and calnexin. This correlates with the absence of the fusion factor N-ethylmaleimide-sensitive factor onto the vacuolar membrane in this cell type. Trafficking between the vacuoles and the host cell recycling and biosynthetic pathways is strikingly reduced in DCs, which is likely to impair access of intracellular mycobacteria to essential nutrients and may thus explain the absence of mycobacterial growth in this cell type. This unique location of M. tuberculosis in DCs is compatible with their T lymphocyte-stimulating functions, because M. tuberculosis-infected DCs have the ability to specifically induce cytokine production by autologous T lymphocytes from presensitized individuals. DCs have evolved unique subcellular trafficking mechanisms to achieve their Ag-presenting functions when infected by intracellular mycobacteria.
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PMID:Constrained intracellular survival of Mycobacterium tuberculosis in human dendritic cells. 1257 62

Fibrillin-1 is a large modular glycoprotein that assembles to form 10-12 nm microfibrils in the extracellular matrix. Mutations in the fibrillin-1 gene (FBN1) cause Marfan syndrome and related connective tissue disorders (fibrillinopathies) that show autosomal dominant inheritance. The pathogenic mechanism is thought to be a dominant negative effect of a mutant protein on microfibril assembly, although direct evidence is lacking. A significant group of disease-causing FBN1 mutations are cysteine substitutions within EGF domains that are predicted to cause misfolding by removal of disulphide bonds that stabilize the native domain fold. We have studied three missense mutations (C1117Y, C1129Y and G1127S) to investigate the effect of misfolding on the trafficking of fibrillin-1 from fibroblast cells. We demonstrate that both C1117Y and C1129Y, expressed as recombinant fragments of fibrillin-1, are retained and accumulate within the cell. Both undergo core glycosylation but lack the complex glycosylation observed in the secreted wild-type fragment, suggesting retention in the endoplasmic reticulum (ER). In addition, co-immunoprecipitation experiments show association with the ER chaperone calreticulin, but not calnexin, 78 kDa glucose-regulated protein (Grp78/BiP) or protein disulfide isomerase. In contrast, G1127S, which causes a moderate change in the EGF domain fold, shows a pattern of glycosylation and trafficking profile indistinguishable from the wild-type fragment. Since expression of the recombinant fragments does not disrupt the secretion of endogenous fibrillin-1 by the cell, we propose that G1127S causes disease via an extracellular dominant negative effect. In contrast, the observed ER retention of C1117Y and C1129Y suggests that disease associated with these missense mutations is caused either by an intracellular dominant negative effect or haploinsufficiency.
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PMID:Defective secretion of recombinant fragments of fibrillin-1: implications of protein misfolding for the pathogenesis of Marfan syndrome and related disorders. 1265 68

In this review we discuss the influence of chaperones on the general phenomena of folding as well as on the specific folding of an individual protein, MHC class I. MHC class I maturation is a highly sophisticated process in which the folding machinery of the endoplasmic reticulum (ER) is heavily involved. Understanding the MHC class I maturation per se is important since peptides loaded onto MHC class I molecules are the base for antigen presentation generating immune responses against virus, intracellular bacteria as well as tumours. This review discusses the early stages of MHC class I maturation regarding BiP and calnexin association, and differences in MHC class I heavy chain (HC) interaction with calnexin and calreticulin are highlighted. Late stage MHC class I maturation with focus on the dedicated chaperone tapasin is also discussed.
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PMID:Chaperones and folding of MHC class I molecules in the endoplasmic reticulum. 1278 24

Mutations in the genes for peripheral myelin protein-22 (PMP22) and myelin protein zero (P0) cause human hereditary neuropathies with varying clinical and pathological phenotypes. In this study, we examine the effects of representative disease-causing mutations on the subcellular distribution of their corresponding PMP22- and P0-enhanced green fluorescent protein (EGFP) fusion proteins. In transiently transfected HeLa and 293 cells, we find that wild-type P0-EGFP and PMP22-EGFP are efficiently synthesized and transported through the secretory pathway to the plasma membrane. The P0-EGFP and PMP22-EGFP mutants can be classified into several groups: those that are transported to the plasma membrane as in the majority of P0 mutants; those that are retained in the endoplasmic reticulum as in the majority of PMP22 mutants; and those that are a mixture of the two. In addition, several of these disease-causing mutations are associated with the development of abnormal intracellular cytoplasmic structures that we have previously identified as either intracellular myelin figures or aggresomes. Our studies indicate that different types of PMP22 and P0 mutations are associated with specific intracellular chaperone proteins, including calnexin and BiP, and that these associations can be altered by glycosylation. These findings indicate that the various P0 and PMP22 mutants may exert their pathogenic effects in different subcellular compartments and by different mechanisms in the mammalian cell.
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PMID:Phenotypic differences between peripheral myelin protein-22 (PMP22) and myelin protein zero (P0) mutations associated with Charcot-Marie-Tooth-related diseases. 1290 1

The limitations to high-level expression of integral membrane proteins are not well understood. The human A(2)a adenosine receptor (A(2)a) and mouse Substance P receptor (SPR) were individually expressed in S. cerevisiae to identify potential cellular bottlenecks for G-protein coupled receptors. In the yeast system, A(2)a was not N-linked glycosylated but was functional and plasma membrane-localized. A(2)a also contained an intramolecular disulfide bond. Substance P receptor was also not N-linked glycosylated in yeast, but, unlike A(2)a, SPR was intracellularly retained, nonfunctional, and did not appear to contain an intramolecular disulfide bond. Since both receptors contain N-linked glycosylation and disulfide bonds in mammalian systems, machinery responsible for interacting with these modifications was investigated-specifically, the potential interactions between the nascent receptor and ER-resident proteins were explored. The chaperones calnexin and protein disulfide isomerase were co-overexpressed with the GPCRs to determine the effect on total and active yields of A(2)a and SPR, as well as on receptor trafficking. The effect of co-expressing the chaperone BiP on the total yields of A(2)a as well as intracellular fates of both receptors were determined. The co-expression of ER resident proteins did not improve A(2)a yields nor did they restore SPR activity or improve SPR cell surface expression. Taken together, these results indicate that an ER-folding bottleneck does not limit the expression of the mammalian receptors in yeast.
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PMID:Co-expression of molecular chaperones does not improve the heterologous expression of mammalian G-protein coupled receptor expression in yeast. 1296 83


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