ERp61 Is GRP58, a Stress-Inducible Luminal Endoplasmic Reticulum Protein, but Is Devoid of Phosphatidylinositide-Specific Phospholipase C Activity

doi:10.1006/abbi.1994.1064

Archives of Biochemistry and Biophysics

Volume 308, Issue 2, February 1994, Pages 454-460

https://doi.org/10.1006/abbi.1994.1064 Get rights and content

Abstract

Using antibody raised against putative Form I phosphatidylinositide-specific phospholipase C (PI-PLC) and direct amino acid sequencing of the protein recognized by this antibody, we have shown that the antibody reacts with luminal endoplasmic reticulum (ER) proteins, including ERp61. ERp61 possesses a COOH-terminal QEDL sequence that acts as an ER retention signal. Additional experiments have shown, however, that PI-PLC activity is separable from ERp61 and that rat or murine ERp61 expressed in COS cells failed to produce an increase in PI-PLC activity in the COS cells. Finally, we have identified ERp61 as GRP58, a 58-kDa protein inducible by glycosylation block and treatment with the Ca²⁺ ionophore, A23187.

References (0)

Cited by (67)

Cerulenin upregulates heat shock protein-70 gene expression in chicken muscle
2013, Poultry Science
Citation Excerpt :
Leptin, the adipocyte-derived hormone, and the key regulator of food intake and energy homeostasis (Zhang et al., 1994) has been shown to modulate key processes involved in stress responses (Bouloumie et al., 1999; Oates et al., 2000). Kita et al. (2006) showed that leptin administration induce phosphorylation of the stress glucose-regulated protein (GRP)-58, which belongs to Hsp families and is induced by glucose starvation and viral infection (Lee, 1987; Mazzarella et al., 1994). We have shown also that recombinant chicken leptin downregulates Hsp-70 gene expression in normal chicken liver and hypothalamus (Figueiredo et al., 2007).
Lines of evidence suggested that systems involved in the regulation of the stress responses and energy homeostasis are highly integrated. Because cerulenin, the natural antibiotic product of the fungus Cephalosporium ceruleans and a broad-spectrum fatty acid synthesis (FAS) inhibitor, has been shown to affect food intake and energy balance, and because the biomarker of stress Hsp-70 gene was found to interact directly with fatty acids, we hypothesized that cerulenin may regulate Hsp-70 gene expression. Therefore, the present study was undertaken to examine this issue. Cerulenin administration significantly (P < 0.05) decreased food intake and induced Hsp-70 mRNA levels in muscle, but not in liver or hypothalamus of 2-wk-old broiler chickens. These changes were accompanied by an unpregulation of muscle uncoupling protein and carnitine palmitoyltransferase 1 mRNA levels. This result indicated that the regulation of Hsp-70 gene expression in normal chickens, as estimated by oxidative stress indices [TBA reacting substances, ferric reducing/antioxidant power, and ceruloplasmin oxidase activity] levels, is tissue-specific. In attempt to discriminate between the effect of cerulenin and cerulenin-reduced food intake on Hsp-70 gene expression, we also evaluated the effect of food deprivation on the same cellular responses. Food deprivation for 16 h did not affect Hsp-70 gene expression in all tissues examined, indicating that the effect of cerulenin is independent of the inhibition of food intake. To ascertain whether the effect of cerulenin is direct or indirect, we carried out in vitro studies. Cerulenin treatment did not affect Hsp-70 gene expression in Leghorn male hepatoma and quail myoblast cell lines, suggesting that the observed effect in vivo may be mediated through the central nervous system.
The cloning and inducible expression of the rainbow trout ERp57 gene
2013, Fish and Shellfish Immunology
Citation Excerpt :
Future studies will determine if ERp57 can be upregulated in virally infected cells as part of the host interferon response. Upon short term induction of ER stress using the calcium ionophore A23187, trout ERp57 protein levels were increased both in RTS11 and PBL, similar to the upregulation of human ERp57 [31] and other ER resident chaperones such as calreticulin [32] and Bip [33] during ER stress. The PCR primers used in the induction studies reported here are specific for the sequence we isolated which lacks the nuclear localization signal, thus may encode the ER specific version of the ERp57 protein.
ERp57 is a member of a protein disulfide isomerase family and is a chaperone responsible for the correct folding of newly synthesized glycoproteins in the endoplasmic reticulum and in the assembly of the major histocompatibility complex class I in the endogenous pathway of antigen presentation. This study reports the identification of a full length ERp57 cDNA in rainbow trout that encodes a putative 477aa mature protein with an additional signal sequence of 16aa. The trout protein shared 75% identity with the human homolog, but interestingly did not include either a C terminal endoplasmic reticulum retention signal, Q/KEDL in humans, or a nuclear localization signal which is highly conserved in mammals. Amino acid sequence alignment revealed conservation of four classical domains in trout ERp57 and two conserved active CXXC redox motifs. Trout ERp57 protein was identified as a single band around 57 kDa. Southern blotting analysis revealed that there two copies of the ERp57 gene in the trout genome and northern blotting showed a wide tissue distribution of gene expression in various tissues with the highest expression in liver and egg. This study showed for the first time in teleost that ERp57 transcript is upregulated in response to immune stimuli such as double stranded RNA or phytohemagglutinin. Furthermore, upon treatment with ER stress inducer A23187, trout ERp57 protein expression levels were increased both in peripheral blood leukocytes and the RTS11 macrophage like cell line after 6 and 8 h respectively. These findings suggest a possible conserved function for trout ERp57 in the ER and during the activation of the immune response.
ERp57 modulates STAT3 signaling from the lumen of the endoplasmic reticulum
2010, Journal of Biological Chemistry
Citation Excerpt :
Interestingly, small interfering RNA studies also demonstrate that ERp57 might be critical for oxidative folding of immunoglobulin heavy chain but not important for peptide loading of class I molecules (5). Aside from its role as a folding enzyme in quality control in the secretory pathway, ERp57, also known as glucose-regulated protein 58 or Grp58 (6–13) has been reported to affect STAT3 (signal transducers and activators of transcription) signaling via interaction with STAT3 (6, 14–17). STATs are a family of cytoplasmic proteins with Src homology 2 domains that act as signal messengers and transcription factors as a part of the Janus kinase-STAT pathway (18).
ERp57 is an endoplasmic reticulum (ER) resident thiol disulfide oxidoreductase. Using the gene trap technique, we created a ERp57-deficient mouse model. Targeted deletion of the Pdia3 gene, which encodes ERp57, in mice is embryonic lethal at embryonic day (E) 13.5. β-Galactosidase reporter gene analysis revealed that ERp57 is expressed early on during blastocyst formation with the highest expression in the inner cell mass. In early stages of mouse embryonic development (E11.5) there is a relatively low level of expression of ERp57. As the embryos developed, ERp57 became highly expressed in both the brain and the lungs (E15.5 and E18.5). The absence of ERp57 has no impact on ER morphology; expression of ER-associated chaperones and folding enzymes, ER stress, or apoptosis. ERp57 has been reported to interact with STAT3 (signal transducer and activator of transcription)-DNA complexes. We show here that STAT3-dependent signaling is increased in the absence of ERp57 and this can be rescued by expression of ER-targeted ERp57 but not by cytoplasmic-targeted protein, indicating that ERp57 affects STAT3 signaling from the lumen of the ER. ERp57 effects on STAT3 signaling are enhanced by ER luminal complex formation between ERp57 and calreticulin. In conclusion, we show that ERp57 deficiency in mouse is embryonic lethal at E13.5 and ERp57-dependent modulation of STAT3 signaling may contribute to this phenotype.
DNA-binding activity of the ERp57 C-terminal domain is related to a redox-dependent conformational change
2007, Journal of Biological Chemistry
ERp57, a member of the protein-disulfide isomerase family, although mainly localized in the endoplasmic reticulum is here shown to have a nuclear distribution. We previously showed the DNA-binding properties of ERp57, its association with the internal nuclear matrix, and identified the C-terminal region, containing the a′ domain, as being directly involved in the DNA-binding activity. In this work, we demonstrate that its DNA-binding properties are strongly dependent on the redox state of the a′ domain active site. Site-directed mutagenesis experiments on the first cysteine residue of the -CGHC-thioredoxin-like active site lead to a mutant domain (C406S) lacking DNA-binding activity. Biochemical studies on the recombinant domain revealed a conformational change associated with the redox-dependent formation of a homodimer, having two disulfide bridges between the cysteine residues of two a′ domain active sites. The formation of intermolecular disulfide bridges rather than intramolecular oxidation of active site cysteines is important to generate species with DNA-binding properties. Thus, in the absence of any dedicated motif within the protein sequence, this structural rearrangement might be responsible for the DNA-binding properties of the C-terminal domain. Moreover, NADH-dependent thioredoxin reductase is active on intermolecular disulfides of the a′ domain, allowing the control of dimeric protein content as well as its DNA-binding activity. A similar behavior was also observed for whole ERp57.
Proteomic analysis of cisplatin-induced cochlear damage: Methods and early changes in protein expression
2007, Hearing Research
To identify early changes in protein expression associated with cisplatin ototoxicity, we used two dimensional-difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption-time-of-flight (MALDI-TOF) mass spectrometry to analyze proteins from P3 rat cochleae that were cultured for 3 h with or without 1 mM cisplatin. Replicate analysis of fluorescent images from six gels revealed significant (p < 0.01) cisplatin-induced changes (greater than 1.5-fold) in expression of 22 cochlear proteins. These include increases in the expression of five proteins, four of which were identified as nucleobindin 1, a nuclear calcium signaling and homeostasis protein (2.1-fold), heterogeneous nuclear ribonucleoprotein C, an RNA processing protein (1.8-fold), a 55 kDa protein that is either endothelial differentiation-related factor 1 or alpha-6 tubulin (1.7-fold), and calreticulin, a calcium binding chaperone of the endoplasmic reticulum (ER, 1.6-fold). The expression of 17 proteins was significantly (p < 0.01) decreased by greater than 1.5-fold. These include ribonuclease/angiogenin inhibitor 1 (1.6-fold), RAS-like, family 12 (predicted), ras association (RalGDS/AF-6) domain family 5 (4.5-fold), homologous the RAS family of GTPase signaling proteins (2.4-fold), and Protein tyrosine phosphatase domain containing 1 (predicted, 6.1-fold). We identified seven cochlear proteins with either smaller (1.2–1.5-fold) or less significant (p < 0.05) cisplatin-induced changes in expression. Notably, heat shock 70 kDa protein 5 (Hspa5, Grp78, and BiP), an ER chaperone protein involved in stress response, decreased 1.7-fold. We observed changes consistent with phosphorylation in the level of isoforms of another ER stress-induced protein, glucose-regulated protein Grp58. Changes in cisplatin-induced protein expression are discussed with respect to known or hypothesized functions of the identified proteins.
Induction of the unfolded protein response in familial amyotrophic lateral sclerosis and association of protein-disulfide isomerase with superoxide dismutase 1
2006, Journal of Biological Chemistry
Citation Excerpt :
We found PDI and Erp57 to be greatly up-regulated in the lumbar spinal cords of SOD1G93A rodents in this study, correlating with the formation of SOD1 aggregates in vivo. Erp57 is a close homologue of PDI (59) whose up-regulation may relate to SOD1 folding (60) or participation in the ER stress response (61). Bacitracin-mediated inhibition of PDI in cell transfectants increased the frequency of mSOD1 inclusions, suggesting that PDI activity plays a role in the prevention of insoluble SOD1 aggregate formation.
Mutations in Cu/Zn superoxide dismutase (SOD1) are linked to motor neuron death in familial amyotrophic lateral sclerosis (ALS) by an unclear mechanism, although misfolded SOD1 aggregates are commonly associated with disease. Proteomic analysis of the transgenic SOD1^G93A ALS rat model revealed significant up-regulation of endoplasmic reticulum (ER)-resident protein-disulfide isomerase (PDI) family members in lumbar spinal cords. Expression of SOD1 mutants (mSOD1) led to an up-regulation of PDI in motor neuron-like NSC-34 cells but not other cell lines. Inhibition of PDI using bacitracin increased aggregate production, even in wild type SOD1 transfectants that do not readily form inclusions, suggesting PDI may protect SOD1 from aggregation. Moreover, PDI co-localized with intracellular aggregates of mSOD1 and bound to both wild type and mSOD1. SOD1 was also found in the microsomal fraction of cells despite being a predominantly cytosolic enzyme, confirming ER-Golgi-dependent secretion. In SOD1^G93A mice, a significant up-regulation of unfolded protein response entities was also observed during disease, including caspase-12, -9, and -3 cleavage. Our findings therefore implicate unfolded protein response and ER stress-induced apoptosis in the patho-physiology of familial ALS. The possibility that PDI may be a therapeutic target to prevent SOD1 aggregation is also raised by this study.

View all citing articles on Scopus

View full text

Regular ArticleERp61 Is GRP58, a Stress-Inducible Luminal Endoplasmic Reticulum Protein, but Is Devoid of Phosphatidylinositide-Specific Phospholipase C Activity

Abstract

Regular Article
ERp61 Is GRP58, a Stress-Inducible Luminal Endoplasmic Reticulum Protein, but Is Devoid of Phosphatidylinositide-Specific Phospholipase C Activity