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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Class I histocompatibility proteins fold and assemble with beta 2-microglobulin (beta 2m) into heterodimers before binding short peptides in the endoplasmic reticulum. Here, we show that class I proteins rapidly form disulfide bonds, and that the process is highly reversible in Daudi cells lacking beta 2m. Three distinct class I protein conformations are present in equal amounts in these cells, each associated with the molecular chaperone calnexin. When binding of calnexin is inhibited by the glucosidase inhibitor castanospermine, fully oxidized class I proteins are no longer detected, suggesting that calnexin is required for completion of folding. However, in Daudi cells transfected to express beta 2m, castanospermine decreases only slightly the levels of fully oxidized class I proteins, indicating that folding is much less dependent on calnexin in the presence of beta 2m. Furthermore, calreticulin, a chaperone with functional similarities to calnexin, associates with class I molecules in beta 2m-positive cells. but not in Daudi cells, consistent with completion of folding and disulfide bond formation of class I heavy chains before binding to calreticulin occurs. This study demonstrates that calnexin and beta 2m can function independently to promote folding of class I heavy chains prior to formation of stable class I dimers.
Mol Immunol 1997 Apr
PMID:Beta 2-microglobulin and calnexin can independently promote folding and disulfide bond formation in class I histocompatibility proteins. 929 73

Calsequestrin (CSQ), the major low-affinity Ca(2+)-binding glycoprotein of striated muscle fibers, is concentrated to yield aggregates that occupy the lumen of the terminal cisternae of the sarcoplasmic reticulum (SR). When infected or transfected into L6 myoblast, the protein is also concentrated, however, in dense vacuoles apparently separate from the endoplasmic reticulum (ER). CSQ-rich cells appear otherwise normal; in particular, neither other proteins involved in Ca2+ homeostasis nor ER chaperones are increased. The CSQ dense vacuoles are shown herein to be specialized ER subdomains as demonstrated by 1) the endoglycosidase H sensitivity of their CSQ and 2) two markers, calreticulin and calnexin (but not others, protein disulfide isomerase and BiP), intermixed with the vacuole content. Their formation is shown to start with the aggregation of CSQ at discrete sites of the ER lumen. When cells were transfected with both CSQ and calreticulin, only the first gave rise to vacuoles; the second remained diffusely distributed within the ER lumen. The possibility that CSQ aggregation is an artifact of overexpression appears unlikely because 1) within dense vacuoles CSQ molecules are not disulfide cross-linked, 2) their turnover is relatively slow (t = 12 h), and 3) segregated CSQ is bound to large amounts of Ca2+. Transfection of a tagged CSQ into cells already overexpressing the protein revealed the continuous import of the newly synthesized protein into preassembled vacuoles. The tendency to aggregation appears, therefore, as a property contributing to the segregation of CSQ within the ER lumen and to its accumulation within specialized subdomains. The study of L6 cells expressing CSQ-rich vacuoles might thus ultimately help to unravel mechanisms by which the complexity of the sarcoplasmic reticulum is established in muscle fibers.
Mol Biol Cell 1997 Sep
PMID:Overexpression of calsequestrin in L6 myoblasts: formation of endoplasmic reticulum subdomains and their evolution into discrete vacuoles where aggregates of the protein are specifically accumulated. 930 74

Unlike properly folded and assembled proteins, most misfolded and incompletely assembled proteins are retained in the endoplasmic reticulum of mammalian cells and degraded without transport to the Golgi complex. To analyze the mechanisms underlying this unique sorting process and its fidelity, the fate of C-terminally truncated fragments of influenza hemagglutinin was determined. An assortment of different fragments was generated by adding puromycin at low concentrations to influenza virus-infected tissue culture cells. Of the fragments generated, < 2% was secreted, indicating that the system for detecting defects in newly synthesized proteins is quite stringent. The majority of secreted species corresponded to folding domains within the viral spike glycoprotein. The retained fragments acquired a partially folded structure with intra-chain disulfide bonds and conformation-dependent antigenic epitopes. They associated with two lectin-like endoplasmic reticulum chaperones (calnexin and calreticulin) but not BiP/GRP78. Inhibition of the association with calnexin and calreticulin by the addition of castanospermine significantly increased fragment secretion. However, it also caused association with BiP/GRP78. These results indicated that the association with calnexin and calreticulin was involved in retaining the fragments. They also suggested that BiP/GRP78 could serve as a backup for calnexin and calreticulin in retaining the fragments. In summary, the results showed that the quality control system in the secretory pathway was efficient and sensitive to folding defects, and that it involved multiple interactions with endoplasmic reticulum chaperones.
Mol Biol Cell 1997 Oct
PMID:Quality control in the secretory pathway: the role of calreticulin, calnexin and BiP in the retention of glycoproteins with C-terminal truncations. 934 35

Calreticulin is a major Ca2+ binding protein in the endoplasmic reticulum of non-muscle cells. In this report we show that calreticulin protein is strongly induced by heat shock. Activation and attenuation of the heat shock transcriptional response is caused by heat shock factor that binds to 5'-flanking sequences of heat shock responsive genes, the heat shock element. The smallest stretch of DNA that shows detectable binding of heat shock factor in vitro contains a two-sequence unit nGAAnnTTCn which exists in the 5'-flanking region of calreticulin DNA (5'-gGAAccCAGcgTTC-3'). The present data provide direct evidence that calreticulin expression can be modulated by heat shock. Thus, our results strengthen the hypothesis that calreticulin, in addition to its function as a cellular Ca2+ store, is a multifunctional protein which performs at least some of its functions from the lumen of the ER.
Mol Cell Biochem 1997 Dec
PMID:Heat shock-regulated expression of calreticulin in retinal pigment epithelium. 945 Jun 56

Since its first identification as a high-affinity calcium-binding protein over two decades ago [T.J. Ostwald and D.H. MacLennan, Isolation of a high-affinity calcium-binding protein from sarcoplasmic reticulum, J. Biol. Chem., 249 (1974) 974-979], calreticulin has become recognized as a multifunctional protein involved in a wide variety of cellular processes. We have previously shown that it has a protective function in Ca2+-mediated cell death [N. Liu, R.E. Fine, E. Simons and R.J. Johnson, Decreasing calreticulin expression lowers the Ca2+ response to bradykinin and increases sensitivity to ionomycin in NG-108-15 cells, J. Biol. Chem. , 269 (1994) 28635-28639]. We report here that in NG-108-15 neuroblastomaxglioma hybrid cells, calreticulin protein levels increase markedly when these cells are induced to differentiate by treating them with N,N-dibutyryl cAMP (db-cAMP). We demonstrate that the reason for this increase is mostly due to a large increase in the turnover time of calreticulin in differentiated cells. We also show that a calreticulin antisense oligonucleotide, CrtAS1, previously described by Liu and co-workers [N. Liu, R.E. Fine, E. Simons and R.J. Johnson, Decreasing calreticulin expression lowers the Ca2+ response to bradykinin and increases sensitivity to ionomycin in NG-108-15 cells, J. Biol. Chem., 269 (1994) 28635-28639] causes cell death in undifferentiated NG-108-15 cells when antisense treatment is extended for more than 24 h. This effect is not seen in NG-108-15 cells that have been induced to differentiate with db-cAMP until the cells have been treated with antisense for more than 4 days, due to the increased stability of Crt in these cells. Our results indicate that the mechanism by which these cells die is likely to be apoptosis.
Brain Res Mol Brain Res 1998 Jan
PMID:Increased calreticulin stability in differentiated NG-108-15 cells correlates with resistance to apoptosis induced by antisense treatment. 947 13

1,25-dihydroxyvitaminD3 [1,25-(OH)2D3] and PTH both act to increase serum calcium. In addition, 1,25-(OH)2D3 decreases PTH gene transcription, which is relevant both to the physiology of calcium homeostasis and to the management of the secondary hyperparathyroidism of patients with chronic renal failure. In chronic hypocalcemia there is secondary hyperparathyroidism with increased levels of PTH mRNA and serum PTH despite markedly increased levels of 1,25-(OH)2D3. We have studied the role of calreticulin in this resistance to 1,25-(OH)2D3. Weanling rats fed a low-calcium diet were hypocalcemic and had increased PTH mRNA levels despite high serum 1,25-(OH)2D3 levels. 1,25-(OH)2D3 given by continuous minipump infusion to normal rats led to the expected decrease in PTH mRNA. The hypocalcemic rats had an increased concentration of calreticulin in the nuclear fraction of their parathyroids, but not in other tissues. Gel shift assays showed that a purified vitamin D receptor and retinoid X receptor-beta bound to the PTH promoter's chicken and rat vitamin D response element (VDRE), and this binding was inhibited by added pure calreticulin. Transfection studies with a PTH VDRE-chloramphenicol acetyltransferase (CAT) construct showed that 1,25-(OH)2D3 decreased CAT transcription. Cotransfection of PTH VDRE-CAT with a calreticulin expression vector in the sense orientation prevented the transcriptional effect of 1,25-(OH)2D3, but a calreticulin vector in the antisense orientation had no effect. These results show that calreticulin prevents the binding of vitamin D receptor-retinoid X receptor-beta to the PTH VDRE in gel retardation assays and prevents the transcriptional effect of 1,25-(OH)2D3 on the PTH gene. This is the first report of calreticulin inhibiting a down-regulatory function of a sterol hormone and may help explain the refractoriness of the secondary hyperparathyroidism of many chronic renal failure patients to 1,25-(OH)2D3.
Mol Endocrinol 1998 Aug
PMID:Calreticulin inhibits vitamin D's action on the PTH gene in vitro and may prevent vitamin D's effect in vivo in hypocalcemic rats. 971 45

A full length D. immitis cDNA (nDiCal) encoding a protein with significant similarity to the calreticulin protein family was isolated from a 6-day fourth-stage larval cDNA expression library by immunoscreening, using serum from a rabbit immunized by repeated injection of small numbers of third-stage larvae. nDiCal is 1538 bp long and contains the 21 bp nematode splice leader sequence SL1 at the 5' end. nDiCal encodes for a protein (pDiCal) with a predicted molecular mass of 46 kDa. pDiCal sequence analysis revealed similarities with calreticulin, a protein that typically resides in the endoplasmic reticulum. pDiCal possesses three consensus sequences of the calreticulin family of proteins: a neutral N-terminal region with a putative signal sequence; a proline- and tryptophan-rich P region; and a highly acidic C-terminal region. A 45Ca2+-overlay assay showed that recombinant pDiCal (rDiCal) is a Ca2+-binding protein. Antibodies to rDiCal identified a 56 kDa native antigen in all developmental stages including the excretory-secretory products derived from larvae and adult worms. Localization studies demonstrated the ubiquitous presence of pDiCal with intense expression in the hypodermis and syncitial muscle cells in both male and female adult worms. Labeling was also seen in the developing embryos within the uterus of the female worms. Sera from immune as well as chronically-infected microfilaremic dogs contained antibodies that bind rDiCal. In addition, immunoblot analysis showed that serum from a rabbit immunized with L3 cuticles reacted with rDiCal.
Mol Biochem Parasitol 1998 Nov 30
PMID:Molecular characterization of a calcium-binding protein from the filarial parasite Dirofilaria immitis. 987 88

Protein kinase CK2 forms complexes with some protein substrates what may be relevant for the physiological control of this protein kinase. In previous studies in rat liver cytosol we had detected that the trimeric form of eukaryotic translation initiation factor 2 (eIF-2) co-eluted with protein kinase CK2. We have now observed that the ratio between eIF-2 and cytosolic CK2 contents in testis, liver and brain is quite similar, being eIF-2 levels about 5-fold higher than those of CK2. Furthermore eIF-2 was present in liver samples immunoprecipitated with anti-CK2alpha/alpha' antibodies, confirming the existence of complexes containing both proteins. Nonetheless, these complexes would represent only a fraction of total cytosolic CK2 and eIF-2. We had also observed that rat liver membrane glycoproteins obtained through chromatography on wheat-germ lectin-Sepharose contain CK2 activity which copurifies with grp94/endoplasmin. We have now confirmed that this activity was due to the presence of protein kinase CK2 as evidenced by immunodetection with antibodies against CK2alpha/alpha'. The fractions enriched in grp94/endoplasmin and CK2 also contained another 55-kDa polypeptide (p55) phosphorylated by CK2 which has been identified as calreticulin by N-terminal sequencing. Calreticulin and grp94/endoplasmin could be partially resolved from CK2 by chromatography on heparin-agarose and almost completely on ConA-Sepharose. However, phosphorylation of immunoprecipitated grp94/endoplasmin was enhanced by its preincubation with purified CK2 prior to immunoprecipitation, what confirms the easy reassociation between these proteins. The association of protein kinase CK2 with eIF-2 and with grp94/endoplasmin may serve to locate the enzyme in the cellular machinery involved in protein synthesis and folding, and reinforces the possible involvement of CK2 in these processes.
Mol Cell Biochem 1999 Jan
PMID:Association of protein kinase CK2 with eukaryotic translation initiation factor eIF-2 and with grp94/endoplasmin. 1009 97

Trypanosoma cruzi is a protozoan parasite that belongs to an early branch in evolution. Although it lacks several features of the pathway of protein N-glycosylation and oligosaccharide processing present in the endoplasmic reticulum of higher eukaryotes, it displays UDP-Glc:glycoprotein glucosyltransferase and glucosidase II activities. It is herewith reported that this protozoan also expresses a calreticulin-like molecule, the third component of the quality control of glycoprotein folding. No calnexin-encoding gene was detected. Recombinant T. cruzi calreticulin specifically recognized free monoglucosylated high-mannose-type oligosaccharides. Addition of anti-calreticulin serum to extracts obtained from cells pulse-chased with [35S]Met plus [35S]Cys immunoprecipitated two proteins that were identified as calreticulin and the lysosomal proteinase cruzipain (a major soluble glycoprotein). The latter but not the former protein disappeared from immunoprecipitates upon chasing cells. Contrary to what happens in mammalian cells, addition of the glucosidase II inhibitor 1-deoxynojirimycin promoted calreticulin-cruzipain interaction. This result is consistent with the known pathway of protein N-glycosylation and oligosaccharide processing occurring in T. cruzi. A treatment of the calreticulin-cruzipain complexes with endo-beta-N-acetylglucosaminidase H either before or after addition of anti-calreticulin serum completely disrupted calreticulin-cruzipain interaction. In addition, mature monoglucosylated but not unglucosylated cruzipain isolated from lysosomes was found to interact with recombinant calreticulin. It was concluded that the quality control of glycoprotein folding appeared early in evolution, and that T. cruzi calreticulin binds monoglucosylated oligosaccharides but not the protein moiety of cruzipain. Furthermore, evidence is presented indicating that glucosyltransferase glucosylated cruzipain at its last folding stages.
Mol Biol Cell 1999 May
PMID:Trypanosoma cruzi calreticulin is a lectin that binds monoglucosylated oligosaccharides but not protein moieties of glycoproteins. 1023 51

ERp57 is a lumenal protein of the endoplasmic reticulum (ER) and a member of the protein disulfide isomerase (PDI) family. In contrast to archetypal PDI, ERp57 interacts specifically with newly synthesized glycoproteins. In this study we demonstrate that ERp57 forms discrete complexes with the ER lectins, calnexin and calreticulin. Specific ERp57/calreticulin complexes exist in canine pancreatic microsomes, as demonstrated by SDS-PAGE after cross-linking, and by native electrophoresis in the absence of cross-linking. After in vitro translation and import into microsomes, radiolabeled ERp57 can be cross-linked to endogenous calreticulin and calnexin while radiolabeled PDI cannot. Likewise, radiolabeled calreticulin is cross-linked to endogenous ERp57 but not PDI. Similar results were obtained in Lec23 cells, which lack the glucosidase I necessary to produce glycoprotein substrates capable of binding to calnexin and calreticulin. This observation indicates that ERp57 interacts with both of the ER lectins in the absence of their glycoprotein substrate. This result was confirmed by a specific interaction between in vitro synthesized calreticulin and ERp57 prepared in solution in the absence of other ER components. We conclude that ERp57 forms complexes with both calnexin and calreticulin and propose that it is these complexes that can specifically modulate glycoprotein folding within the ER lumen.
Mol Biol Cell 1999 Aug
PMID:ERp57 functions as a subunit of specific complexes formed with the ER lectins calreticulin and calnexin. 1043 13


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