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At least 19 recordsLinked to original sources
PubMed2002

Early defect in the expression of mouse sperm DNAJ 1, a member of the DNAJ/heat shock protein 40 chaperone protein family, in the spinal cord of the wobbler mouse, a murine model of motoneuronal degeneration.

Prevention of protein misfolding is ensured by chaperone proteins, including the heat shock proteins (HSP) of the DNAJ/HSP40 family. Detection of abnormal protein aggregates in various neurodegenerative diseases has led to the proposal that altered chaperone activity contributes to neurodegeneration. Msj-1, a DNAJ/HSP40 protein located around the spermatozoa acrosome, was recently found to be down-regulated in the testis of wobbler mutant mice. Wobbler is an unidentified recessive mutation which triggers progressive motoneuron degeneration with abnormal intracellular protein accumulations, and defective spermatozoa maturation. Here, we examined Msj-1 expression in the spinal cord of the mutants and their controls. Msj-1 transcripts were amplified by reverse transcription-polymerase chain reaction from mutant and wild-type spinal cord RNA. Sequencing of Msj-1 coding region revealed no change in the mutant. In contrast, decreased Msj-1 mRNA levels were observed in five to six-week-old wobbler mice spinal cord, when motoneuron degeneration is at its apex, as compared to controls. A similar decrease was observed in two-week-old wobbler spinal cord, when the number of motoneurons is still unaltered, indicating that the decreased mRNA content is intrinsic to the mutant and not simply related to the loss of cells expressing Msj-1. Assays of Msj-1 protein levels yielded similar results. Immunofluorescent labeling revealed numerous Msj-1-ir motoneurons in five-week-old control spinal cord while no signal was observed in age-matched wobbler. Our results show, therefore, that Msj-1 expression is down-regulated in both organs affected by the wobbler mutation, the CNS and the testis, and that this defect precedes the first histological signs of motoneuron degeneration. These results provide the first example of an association between transcriptional repression of a chaperone protein and a neurodegenerative process.

Animals↗
PubMed1994

Protein chaperones and protein folding.

A universal strategy for obtaining maximal protein expression or refolding remains elusive; however, headway has been made toward understanding these processes in vivo. The observation of reversible protein aggregation, asymmetry in protein-chaperone complexes, redox effects on disulfide formation, and the sequential involvement of multiple chaperones and foldases may suggest new approaches. Such new approaches include immobilized catalysts and manipulation of the bacterial periplasm.

Animals↗
PubMed1988

Homologous plant and bacterial proteins chaperone oligomeric protein assembly.

An abundant chloroplast protein is implicated in the assembly of the oligomeric enzyme ribulose bisphosphate carboxylase-oxygenase, which catalyses photosynthetic CO2-fixation in higher plants. The product of the Escherichia coli groEL gene is essential for cell viability and is required for the assembly of bacteriophage capsids. Sequencing of the groEL gene and the complementary cDNA encoding the chloroplast protein has revealed that these proteins are evolutionary homologues which we term 'chaperonins'. Chaperonins comprise a class of molecular chaperones that are found in chloroplasts, mitochondria and prokaryotes. Assisted post-translational assembly of oligomeric protein structures is emerging as a general cellular phenomenon.

Adenosine Triphosphatases↗
PubMed2001

Expression patterns of chaperone proteins in cerebral cortex of the fetus with Down syndrome: dysregulation of T-complex protein 1.

Chaperone proteins reduce stress-induced denaturation and aggregation of intracellular proteins and exert protective actions by interfering with the stress-induced apoptotic pathway in the neuronal cells. Although many studies have focused on the critical role of chaperones in protein folding and their relevance in protein conformational diseases such as Alzheimer's disease (AD) and prion disease, limited information is available on their expression pattern in AD-related disease, Down Syndrome (DS). In a previous study, we reported differentially expressed molecular chaperone proteins in the brains of adult DS patients. This report demonstrated the relevance of the aberrant expression patterns to biochemical and neuropathological abnormalities in adult DS brain. To investigate whether this aberrant expression patterns already exist in second trimester DS brain, we performed two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix-associated laser desorption ionization mass spectroscopy (MALDI-MS) using fetal DS brain. We unambiguously identified and quantified twelve heat shock proteins (HSPs), three glucose regulated proteins (GRPs) and eight T-complex 1 (TCP-1) subunits. Unlike in adult brain with DS, these studies provide evidence for the normal response of most molecular chaperone proteins at the given time point of DS brain development. However, chaperonin protein TCP-1 alpha and beta subunits showed a significant decrease (P < 0.05) in second trimester brain with DS. Comparable response of chaperone proteins in fetal DS brain suggests that not heat-shock induced proteins but rather chaperonin protein TCP-1 may contribute significantly to neuropathogenesis in the early second trimester DS brain. This study extends observations of distinct expression patterns of molecular chaperones in DS brain and we discuss the relevance of aberrant TCP-1 expression patterns to developmental DS brain at the early stage.

Cerebral Cortex↗
PubMed1998

NMR solution structure of the 21 kDa chaperone protein DnaK substrate binding domain: a preview of chaperone-protein interaction.

The solution structure of the 21 kDa substrate-binding domain of the Escherichia coli Hsp70-chaperone protein DnaK (DnaK 386-561) has been determined to a precision of 1.00 A (backbone of the beta-domain) from 1075 experimental restraints obtained from multinuclear, multidimensional NMR experiments. The domain is observed to bind to its own C-terminus and offers a preview of the interaction of this chaperone with other proteins. The bound protein region is tightly held at a single amino acid position (a leucyl residue) that is buried in a deep pocket lined with conserved hydrophobic residues. A second hydrophobic binding site was identified using paramagnetically labeled peptides. It is located in a region close to the N-terminus of the domain and may constitute the allosteric region that links substrate-binding affinity with nucleotide binding in the Hsp70 chaperones.

Amino Acid Sequence↗
PubMed1993

Highly selective binding of nascent polypeptides by an Escherichia coli chaperone protein in vivo.

Chaperone proteins bind to newly synthesized polypeptides and assist in various assembly reactions. The Escherichia coli chaperone protein SecB binds precursors of exported proteins and assists in export. In vitro, SecB can bind to many unfolded proteins. In this report, we demonstrate that SecB binding in vivo is highly selective; the major polypeptides that are bound by SecB are nascent precursors of the exported proteins maltose-binding protein (MBP), LamB, OmpF, and OmpA. These results support the hypothesis that the primary physiological function of SecB is to stimulate protein export. By interacting with nascent polypeptides, SecB probably stimulates their cotranslational association with the membrane-bound protein translocation apparatus.

Bacterial Proteins↗
PubMed2003

Between genotype and phenotype: protein chaperones and evolvability.

Protein chaperones direct the folding of polypeptides into functional proteins, facilitate developmental signalling and, as heat-shock proteins (HSPs), can be indispensable for survival in unpredictable environments. Recent work shows that the main HSP chaperone families also buffer phenotypic variation. Chaperones can do this either directly through masking the phenotypic effects of mutant polypeptides by allowing their correct folding, or indirectly through buffering the expression of morphogenic variation in threshold traits by regulating signal transduction. Environmentally sensitive chaperone functions in protein folding and signal transduction have different potential consequences for the evolution of populations and lineages under selection in changing environments.

Animals↗
PubMed2002

The ubiquitous nature of RNA chaperone proteins.

RNA chaperones are ubiquitous and abundant proteins found in all living organisms and viruses, where they interact with various classes of RNA. These highly diverse families of nucleic acid-binding proteins possess activities enabling rapid and faithful RNA-RNA annealing, strand transfer, and exchange and RNA ribozyme-mediated cleavage under physiological conditions. RNA chaperones appear to be critical to functions as important as maintenance of chromosome ends, DNA transcription, preRNA export, splicing and modifications, and mRNA translation and degradation. Here we review some of the properties of RNA chaperones in RNA-RNA interactions that take place during cellular processes and retrovirus replication. Examples of cellular and viral proteins are dicussed vis à vis the relationships between RNA chaperone activities in vitro and functions. In this new "genomic era" we discuss the possible use of small RNA chaperones to improve the synthesis of cDNA libraries for use in large screening reactions using DNA chips.

Animals↗
PubMed1998

Proparathyroid hormone-related protein is associated with the chaperone protein BiP and undergoes proteasome-mediated degradation.

Parathyroid hormone-related peptide (PTHrP) is an important causal factor for hypercalcemia associated with malignancy. In addition to the endocrine functions attributed to secretory forms of the peptide, PTHrP also plays a local role as a mediator of cellular growth and differentiation presumably at least in part through intracellular pathways. In studying the post-translational regulation of PTHrP, we observed that PTHrP was conjugated to multiple ubiquitin moieties. We report here that the proteasome is responsible for the degradation of the endoplasmic reticulum-associated precursor, pro-PTHrP. Cells expressing prepro-PTHrP and exposed to lactacystin accumulate pro-PTHrP assessed by anti-pro specific antibodies. Brefeldin A-treated cells also accumulate pro-PTHrP suggesting that degradation does not occur in the endoplasmic reticulum (ER) lumen. Subcellular fractionation of both lactacystin and brefeldin A-treated cells indicated that accumulated pro-PTHrP resides in microsomal fractions with a portion of the protein exposed to the cytosolic side of the ER membrane as assessed by protease protection experiments. Immunoprecipitation and Western blot analysis identified pro-PTHrP in association with the ER molecular chaperone protein BiP. We conclude that pro-PTHrP from the ER can gain access to the cytoplasmic side of the ER membrane where it can undergo ubiquitination and degradation by the proteasome.

Acetylcysteine↗
PubMed2005

Interactions among yeast protein-disulfide isomerase proteins and endoplasmic reticulum chaperone proteins influence their activities.

We previously reported that the reductive activities of yeast protein-disulfide isomerase (PDI) family proteins did not completely explain their contribution to the viability of Saccharomyces cerevisiae (Kimura, T., Hosoda, Y., Kitamura, Y., Nakamura, H., Horibe, T., and Kikuchi, M. (2004) Biochem. Biophys. Res. Commun. 320, 359-365). In this study, we examined oxidative refolding activities and found that Mpd1p, Mpd2, and Eug1p exhibit activities of 13.8, 16.0, and 2.16%, respectively, compared with Pdi1p and that activity for Eps1p is undetectable. In analyses of interactions between yeast PDI proteins and endoplasmic reticulum molecular chaperones, we found that Mpd1p alone does not have chaperone activity but that it interacts with and inhibits the chaperone activity of Cne1p, a homologue of mammalian calnexin, and that Cne1p increases the reductive activity of Mpd1p. These results suggest that the interface between Mpd1p and Cne1p is near the peptide-binding site of Cne1p. In addition, Eps1p interacts with Pdi1p, Eug1p, Mpd1p, and Kar2p with dissociation constants (KD) in the range of 10(-7) to 10(-6). Interestingly, co-chaperone activities were completely suppressed in Eps1p-Pdi1p and Eps1p-Mpd1p complexes, although only Eps1p and Pdi1p have chaperone activity. The in vivo consequences of these results are discussed.

Calnexin↗
PubMed2000

Protein kinase C [micro] is regulated by the multifunctional chaperon protein p32.

We identified the multifunctional chaperon protein p32 as a protein kinase C (PKC)-binding protein interacting with PKCalpha, PKCzeta, PKCdelta, and PKC mu. We have analyzed the interaction of PKC mu with p32 in detail, and we show here in vivo association of PKC mu, as revealed from yeast two-hybrid analysis, precipitation assays using glutathione S-transferase fusion proteins, and reciprocal coimmunoprecipitation. In SKW 6.4 cells, PKC mu is constitutively associated with p32 at mitochondrial membranes, evident from colocalization with cytochrome c. p32 interacts with PKC mu in a compartment-specific manner, as it can be coimmunoprecipitated mainly from the particulate and not from the soluble fraction, despite the presence of p32 in both fractions. Although p32 binds to the kinase domain of PKC mu, it does not serve as a substrate. Interestingly, PKC mu-p32 immunocomplexes precipitated from the particulate fraction of two distinct cell lines, SKW 6.4 and 293T, show no detectable substrate phosphorylation. In support of a kinase regulatory function of p32, addition of p32 to in vitro kinase assays blocked, in a dose-dependent manner, aldolase but not autophosphorylation of PKC mu, suggesting a steric hindrance of substrate within the kinase domain. Together, these findings identify p32 as a novel, compartment-specific regulator of PKC mu kinase activity.

Animals↗
PubMed2004

Complex dynamics of chaperone-protein interactions under cellular stress.

We present a model of a generalizable but minimalistic network based on the properties of interactions between proteins, molecular chaperones (e.g., Hsp70, BiP) and ATP inside cells and subcellular components such as endoplasmic reticulum (ER). The dynamics of chaperone-dependent protein folding and misfolding in the cell can be modeled mathematically as a "predator-prey" problem, which can then be used to analyze the behavior of the system under conditions simulating stress (e.g., cardiac ischemia). We have tested this model under normal physiological and diseased conditions (e.g., ischemia as simulated by ATP depletion) and analyzed the effects of induction of chaperones (e.g., heat shock, tunicamycin) and inhibition of the degradative pathway (e.g., proteasome inhibition) on this model. Simulation gave the following results: (1) Under normal physiological conditions, as expected, the model predicts the stable production of correctly folded proteins. (2) A threshold of ATP levels exists below which the system tends toward increasing degrees of complex behavior. When ATP levels are just above this threshold, the system is highly vulnerable to sudden, brief drops in ATP levels such as may occur in the setting of acute ischemia: bursts of oscillations continue even when ATP levels revert to the threshold. However, if ATP levels are rapidly increased to levels considerably above the threshold, the system becomes stable again. (3) Up to 10-fold increases in chaperone levels, such as those that occur under conditions of prior heat shock or tunicamycin treatment, did not affect the behavior of the system under basal conditions, nor did it affect the tendency to complex behavior in the setting of ATP depletion. It did, however, shorten the recovery period of the system after chaotic-type oscillations were induced by acute ATP depletion. (4) Blocking the degradative pathway for misfolded proteins (e.g., proteasome inhibition) predisposes the system toward instability in the setting of ATP depletion by changing the ATP threshold at which bursts of oscillations occur. These results support the hypothesis that there are distinct thresholds for ATP, chaperones, and degradative activity, outside which cellular protein folding dynamics become unstable. They also suggest that an important mechanism by which chaperone induction protects cells from subsequent stress is by limiting the tendency to instability after an insult (e.g., acute myocardial ischemia or acute tubular injury to the kidney).

Adenosine Triphosphate↗
PubMed1998

Chaperone protein GrpE and the GroEL/GroES complex promote the correct folding of tobacco mosaic virus coat protein for ribonucleocapsid assembly in vivo.

Several prokaryotic chaperone proteins were shown to promote the correct folding and in vivo assembly of tobacco mosaic virus coat protein (TMV CP) using a chimaeric RNA packaging system in control or chaperone-deficient mutant strains of Escherichia coli. Mutations in groEL or dnaK reduced the amount of both total and soluble TMV CP, and the yield of assembled TMV-like particles, several-fold. Thus both GroEL and DnaK have significant direct or indirect effects on the overall expression, stability, folding and assembly of TMV CP in vivo. In contrast, while cells carrying a mutation in grpE expressed TMV CP to a higher overall level than control E. coli, the amounts of both soluble CP and assembled TMV-like particles were below control levels, suggesting a negative effect of GrpE on overall CP accumulation, but positive role(s) in CP folding and assembly. Curiously, cells with mutations in groES and, to a lesser extent, dnaJ expressed total, soluble and assembled forms of TMV CP significantly above control values, suggesting some form of negative control by these chaperone proteins. To avoid pleiotropic effects or artefacts in chaperone-null mutants, selected chaperone proteins were also over-expressed in control E. coli cells. Overproduction of GroEL or GroES alone had little effect. However, co-overexpression of GroEL and GroES resulted in a two-fold increase in soluble TMV CP and a four-fold rise in assembled TMV-like (pseudovirus) particles in vivo. Moreover, TMV CP was shown to interact directly with GroEL in vivo. Together, these results suggest that GrpE and the GroEL/GroES chaperone complex promote the correct folding and assembly of TMV CP into ribonucleocapsids in vivo.

Bacterial Proteins↗
PubMed2004

Regulation of nucleocytoplasmic localizatioin of the atDjC6 chaperone protein.

The sequence of the atDjC6 chaperone protein includes three potential nuclear localization signal (NLS) sequences (A-C) and three potential nuclear export signal (NES) sequences (X-Z). The subcellular localization of atDjC6 was studied by scanning laser confocal microscopy of chimera with the green-fluorescent protein (GFP) transiently expressed in tobacco BY-2 cells. The localization of the atDjC6::GFP chimera was coincident with that of the nuclear stain propidium iodide. Site-directed mutagenesis was used to verify the predicted NLS sequences. Each was individually fused to GFP and tested for protein localization. The individual NLS sequences were sufficient to direct partial nuclear localization of GFP, although the targeting information within NLS-B is apparently conformation sensitive. Site-directed mutagenesis of the NES sequences increased the amount of each chimera that was nuclearly localized, indicating a decrease in nuclear export. When any pair of NLS sequences were appended to GFP, the chimera were entirely nuclearly localized. Quantitative two-hybrid analysis was used to verify that the decoding of NLS sequence information involves interaction with the NLS-receptor protein importin-alpha. Each of the NLS sequences is flanked by a site of potential Ser phosphorylation, and recombinant atDjC6 could be phosphorylated in vitro. Mutagenesis of Ser residues to the P-Ser mimic Asp interfered with nuclear targeting, apparently by preventing recognition or binding by importin-alpha. Our results are consistent with a regulated nucleocytoplasmic localization of the atDjC6 chaperone protein.

Algorithms↗
PubMed2005

Chaperone proteins and brain tumors: potential targets and possible therapeutics.

Chaperone proteins are most notable for the proteo- and cyotoprotective capacities they afford during cellular stress. Under conditions of cellular normalcy, chaperones still play integral roles in the folding of nascent polypeptides into functional entities, in assisting in intracellular/intraorganellar transport, in assembly and maintenance of multi-subunit protein complexes, and in aiding and abetting the degradation of senescent proteins. Tumors frequently have relatively enhanced needs for chaperone number and activity because of the stresses of rapid proliferation, increased metabolism, and overall genetic instability. Thus, it may be possible to take advantage of this reliance that tumor cells have on chaperones by pharmacologic and biologic means. Certain chaperones are abundant in the brain, which implies important roles for them. While it is presumed that the requirements of brain tumors for chaperone proteins are similar to those of any other cell type, tumor or otherwise, very little inquiry has been directed at the possibility of using chaperone proteins as therapeutic targets or even as therapeutic agents against central nervous system malignancies. This review highlights some of the research on the functions of chaperone proteins, on what can be done to modify those functions, and on the physiological responses that tumors and organisms can have to chaperone-targeted or chaperone-based therapies. In particular, this review will also underscore areas of research where brain tumors have been part of the field, although in general those instances are few and far between. This relative dearth of research devoted to chaperone protein targets and therapeutics in brain tumors reveals much untrodden turf to explore for potential treatments of these dreadfully refractive diseases.

Animals↗
PubMed1989

Three pure chaperone proteins of Escherichia coli--SecB, trigger factor and GroEL--form soluble complexes with precursor proteins in vitro.

Diverse studies of three cytoplasmic proteins of Escherichia coli--SecB, trigger factor and GroEL--have suggested that they can maintain precursor proteins in a conformation which is competent for membrane translocation. These proteins have been termed 'chaperones'. Using purified chaperone proteins and precursor protein substrates, we find that each of these chaperones can stabilize proOmpA for translocation and for the translocation-ATPase. These chaperones bind to proOmpA to form isolable complexes. SecB and GroEL will also form complexes with another exported protein, prePhoE. In contrast, these chaperones do not form stable complexes with a variety of soluble proteins such as SecA protein, bovine serum albumin, ovalbumin or ribonuclease A. While chaperones may transiently interact with soluble proteins to catalyze their folding, the stable interaction between chaperones and presecretory proteins, maintaining an open conformation which is essential for translocation, may commit these proteins to the secretion pathway.

Adenosine Triphosphatases↗
PubMed2001

Calreticulin, PDI, Grp94 and BiP chaperone proteins are associated with retained COMP in pseudoachondroplasia chondrocytes.

Cartilage oligomeric matrix protein (COMP), a large pentameric glycoprotein and member of the thrombospondin (TSP) group of extracellular proteins, is found in the territorial matrix surrounding chondrocytes. More than 50 unique COMP mutations have been identified as causing two skeletal dysplasias: pseudoachondroplasia (PSACH); and multiple epiphyseal dysplasia (EDM1). Recent studies suggest that calcium-binding and calcium-induced protein folding differ between wild type and mutant proteins, and abnormal processing of the mutant COMP protein contributes to the characteristic enlarged lamellar appearing rER cisternae in PSACH and EDMI chondrocytes in vivo and in vitro. Towards the goal of delineating the pathogenesis of PSACH and EDM1, in-vivo PSACH growth plate and in-vitro PSACH chondrocytes cultured in alginate beads were examined to identify and localize the chaperone proteins participating in the processing of the retained extracellular matrix proteins in the PSACH rER. Aggrecan was localized to both the rER cisternae and matrix while COMP and type IX collagen were only found in the rER. Type II collagen was solely found in the ECM suggesting that it is processed and transported differently from other retained ECM proteins. Five chaperone proteins: BiP (Grp78); calreticulin (CRT); protein disulfide (PDI); ERp72; and Grp94, demonstrated immunoreactivity in the enlarged PSACH cisternae and the short rER channels of chondrocytes from both in-vivo and in-vitro samples. The chaperone proteins cluster around the electron dense material within the enlarged rER cisternae. CRT, PDI and GRP94 AB-gold particles appear to be closely associated with COMP. Immunoprecipitation and Western blot, and Fluorescence Resonance Energy Transfer (FRET) analyses indicate that CRT, PDI and GRP94 are in close proximity to normal and mutant COMP and BiP to mutant COMP. These results suggest that these proteins play a role in the processing and transport of wild type COMP in normal chondrocytes and in the retention of mutant COMP in PSACH chondrocytes.

Achondroplasia↗
PubMed2003

Identification of alkylation-sensitive target chaperone proteins and their reactivity with natural products containing Michael acceptor.

Molecular chaperones have a crucial role in the folding of nascent polypeptides in endoplasmic reticulum. Some of them are known to be sensitive to the modification by electrophilic metabolites of organic pro-toxicants. In order to identify chaperone proteins sensitive to alkyators, ER extract was subjected to alkylation by 4-acetamido-4'-maleimidyl-stilbene-2,2'-disulfonate (AMS), and subsequent SDS-PAGE analyses. Protein spots, with molecular mass of 160, 100, 57 and 36 kDa, were found to be sensitive to AMS alkylation, and one abundant chaperon protein was identified to be protein disulfide isomerase (PDI) in comparison with the purified PDI. To see the reactivity of PDI with cysteine alkylators, the reduced form (PDIred) of PDI was incubated with various alkylators containing Michael acceptor structure for 30 min at 38 degrees C at pH 6.3, and the remaining activity was determined by the insulin reduction assay. Iodoacetamide or N-ethylmaleimide at 0.1 mM remarkably inactivated PDIred with N-ethylmaleimide being more potent than iodoacetamide. A partial inactivation of PDIoxid was expressed by iodoacetamide, but not N-ethylmaleimide (NEM) at pH 6.3. Of Michael acceptor compounds tested, 1,4-benzoquinone (IC50, 15 microM) was the most potent, followed by 4-hydroxy-2-nonenal and 1,4-naphthoquinone. In contrast, 1,2-naphthoquinone, devoid of a remarkable inactivation action, was effective to cause the oxidative conversion of PDIred to PDIoxid. Thus, the action of Michael acceptor compounds differed greatly depending on their structure. Based on these, it is proposed that PDI, one of chaperone proteins in ER, could be susceptible to endogenous or xenobiotic Michael acceptor compounds in vivo system.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗