scholarly journals The yeast FIT2 homologs are necessary to maintain cellular proteostasis and membrane lipid homeostasis

2020 ◽  
Vol 133 (21) ◽  
pp. jcs248526 ◽  
Author(s):  
Wei Sheng Yap ◽  
Peter Shyu ◽  
Maria Laura Gaspar ◽  
Stephen A. Jesch ◽  
Charlie Marvalim ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Endoplasmic Reticulum ◽  
Lipid Droplets ◽  
Membrane Lipid ◽  
Model System ◽  
Lipid Homeostasis ◽  
Compensatory Mechanism ◽  
Unfolded Protein ◽  
Protein Response

ABSTRACTLipid droplets (LDs) are implicated in conditions of lipid and protein dysregulation. The fat storage-inducing transmembrane (FIT; also known as FITM) family induces LD formation. Here, we establish a model system to study the role of the Saccharomyces cerevisiae FIT homologues (ScFIT), SCS3 and YFT2, in the proteostasis and stress response pathways. While LD biogenesis and basal endoplasmic reticulum (ER) stress-induced unfolded protein response (UPR) remain unaltered in ScFIT mutants, SCS3 was found to be essential for proper stress-induced UPR activation and for viability in the absence of the sole yeast UPR transducer IRE1. Owing to not having a functional UPR, cells with mutated SCS3 exhibited an accumulation of triacylglycerol within the ER along with aberrant LD morphology, suggesting that there is a UPR-dependent compensatory mechanism that acts to mitigate lack of SCS3. Additionally, SCS3 was necessary to maintain phospholipid homeostasis. Strikingly, global protein ubiquitylation and the turnover of both ER and cytoplasmic misfolded proteins is impaired in ScFITΔ cells, while a screen for interacting partners of Scs3 identifies components of the proteostatic machinery as putative targets. Together, our data support a model where ScFITs play an important role in lipid metabolism and proteostasis beyond their defined roles in LD biogenesis.This article has an associated First Person interview with the first author of the paper.

scholarly journals Yeast FIT2 homolog is necessary to maintain cellular proteostasis by regulating lipid homeostasis

2020 ◽  
Author(s):  
Peter Shyu ◽  
Wei Sheng Yap ◽  
Maria L. Gaspar ◽  
Stephen A. Jesch ◽  
Charlie Marvalim ◽  
...  
Keyword(s):  
Stress Response ◽  
Lipid Homeostasis ◽  
Compensatory Mechanism ◽  
Intracellular Lipids ◽  
Cellular Processes ◽  
Unfolded Protein ◽  
Protein Ubiquitination ◽  
Evolutionarily Conserved ◽  
Protein Response

Lipid droplets (LDs) have long been regarded as inert cytoplasmic organelles with the primary function of housing excess intracellular lipids. More recently, LDs have been strongly implicated in conditions of lipid and protein dysregulation. The fat storage inducing transmembrane (FIT) family of proteins comprises of evolutionarily conserved endoplasmic reticulum (ER)-resident proteins that have been reported to induce LD formation. Here, we establish a model system to study the role of S. cerevisiae FIT homologues (ScFIT), SCS3 and YFT2, in proteostasis and stress response pathways. While LD biogenesis and basal ER stress-induced unfolded protein response (UPR) remain unaltered in ScFIT mutants, SCS3 was found to be essential for proper stress-induced UPR activation and for viability in the absence of the sole yeast UPR transducer IRE1. Devoid of a functional UPR, scs3 mutants exhibited accumulation of triacylglycerol within the ER along with aberrant LD morphology, suggesting a UPR-dependent compensatory mechanism for LD maturation. Additionally, SCS3 was necessary to maintain phospholipid homeostasis. Strikingly, the absence of the ScFIT proteins results in the downregulation of the closely-related Heat Shock Response (HSR) pathway. In line with this observation, global protein ubiquitination and the turnover of both ER and cytoplasmic misfolded proteins is impaired in ScFIT cells, while a screen for interacting partners of Scs3 identifies components of the proteostatic machinery as putative targets. Taken together, these suggest that ScFIT proteins may modulate proteostasis and stress response pathways with lipid metabolism at the interface between the two cellular processes.

scholarly journals A Role for the DnaJ Homologue Scj1p in Protein Folding in the Yeast Endoplasmic Reticulum

1998 ◽  
Vol 143 (4) ◽  
pp. 921-933 ◽  
Author(s):  
Susana Silberstein ◽  
Gabriel Schlenstedt ◽  
Pam A. Silver ◽  
Reid Gilmore
Keyword(s):  
Endoplasmic Reticulum ◽  
Unfolded Protein Response ◽  
Physiological Role ◽  
Carboxypeptidase Y ◽  
Reporter Protein ◽  
Unfolded Protein ◽  
A Cell ◽  
The Unfolded Protein Response ◽  
Protein Response

Members of the eukaryotic heat shock protein 70 family (Hsp70s) are regulated by protein cofactors that contain domains homologous to bacterial DnaJ. Of the three DnaJ homologues in the yeast rough endoplasmic reticulum (RER; Scj1p, Sec63p, and Jem1p), Scj1p is most closely related to DnaJ, hence it is a probable cofactor for Kar2p, the major Hsp70 in the yeast RER. However, the physiological role of Scj1p has remained obscure due to the lack of an obvious defect in Kar2p-mediated pathways in scj1 null mutants. Here, we show that the Δscj1 mutant is hypersensitive to tunicamycin or mutations that reduce N-linked glycosylation of proteins. Although maturation of glycosylated carboxypeptidase Y occurs with wild-type kinetics in Δscj1 cells, the transport rate for an unglycosylated mutant carboxypeptidase Y (CPY) is markedly reduced. Loss of Scj1p induces the unfolded protein response pathway, and results in a cell wall defect when combined with an oligosaccharyltransferase mutation. The combined loss of both Scj1p and Jem1p exaggerates the sensitivity to hypoglycosylation stress, leads to further induction of the unfolded protein response pathway, and drastically delays maturation of an unglycosylated reporter protein in the RER. We propose that the major role for Scj1p is to cooperate with Kar2p to mediate maturation of proteins in the RER lumen.

scholarly journals XBP-1 Regulates a Subset of Endoplasmic Reticulum Resident Chaperone Genes in the Unfolded Protein Response

2003 ◽  
Vol 23 (21) ◽  
pp. 7448-7459 ◽  
Author(s):  
Ann-Hwee Lee ◽  
Neal N. Iwakoshi ◽  
Laurie H. Glimcher
Keyword(s):  
Endoplasmic Reticulum ◽  
Unfolded Protein Response ◽  
Target Genes ◽  
Gene Profiling ◽  
Minor Role ◽  
Compensatory Mechanism ◽  
Unfolded Protein ◽  
The Unfolded Protein Response ◽  
A Minor ◽  
Protein Response

ABSTRACT The mammalian unfolded protein response (UPR) protects the cell against the stress of misfolded proteins in the endoplasmic reticulum (ER). We have investigated here the contribution of the UPR transcription factors XBP-1, ATF6α, and ATF6β to UPR target gene expression. Gene profiling of cell lines lacking these factors yielded several XBP-1-dependent UPR target genes, all of which appear to act in the ER. These included the DnaJ/Hsp40-like genes, p58IPK, ERdj4, and HEDJ, as well as EDEM, protein disulfide isomerase-P5, and ribosome-associated membrane protein 4 (RAMP4), whereas expression of BiP was only modestly dependent on XBP-1. Surprisingly, given previous reports that enforced expression of ATF6α induced a subset of UPR target genes, cells deficient in ATF6α, ATF6β, or both had minimal defects in upregulating UPR target genes by gene profiling analysis, suggesting the presence of compensatory mechanism(s) for ATF6 in the UPR. Since cells lacking both XBP-1 and ATF6α had significantly impaired induction of select UPR target genes and ERSE reporter activation, XBP-1 and ATF6α may serve partially redundant functions. No UPR target genes that required ATF6β were identified, nor, in contrast to XBP-1 and ATF6α, did the activity of the UPRE or ERSE promoters require ATF6β, suggesting a minor role for it during the UPR. Collectively, these results suggest that the IRE1/XBP-1 pathway is required for efficient protein folding, maturation, and degradation in the ER and imply the existence of subsets of UPR target genes as defined by their dependence on XBP-1. Further, our observations suggest the existence of additional, as-yet-unknown, key regulators of the UPR.

scholarly journals Nox4 Mediates RANKL-induced ER-Phagy and Osteoclastogenesis via Activating ROS/PERK/eIF-2α/ATF4 Pathway

2020 ◽  
Author(s):  
Jing Sun ◽  
wugui chen ◽  
Songtao Li ◽  
Sizhen Yang ◽  
Ying Zhang ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Bone Resorption ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Nuclear Factor Κb ◽  
Protein Levels ◽  
Unfolded Protein ◽  
Regulatory Effects ◽  
Protein Response

Abstract Background: Receptor activator of nuclear factor-κB ligand (RANKL) has been found to induce osteoclastogenesis and bone resorption. However, the underlying molecular mechanisms remain unclear. Methods: Osteoclastogenesis was evaluated by number of TRAP-positive multinuclear (≥3) osteoclasts, bone resorption pits and expression levels of related genes. Autophagy activity were evaluated by LC3-II/LC3-I ratio, number of autophagic vacuoles and adenovirus-mRFP-GFP-tagged LC3 reporting system; Inhibitor chloroquine (CQ) was used to verified the role of autophagy in RANKL-induced osteoclastogenesis; Via downregulating Nox4 with inhibitor (DPI) and retrovirus-conveyed shRNA, we further explored the importance of Nox4 in RANKL-induced autophagy and osteoclastogenesis, as well as the regulatory effects of Nox4 on nonmitochondrial reactive oxygen species (ROS) and PERK/eIF-2α/ATF4 pathway. Intracellular ROS scavenger (NAC), mitochondrial-targeted antioxidant (MitoTEMPO) and inhibitor of PERK (GSK2606414) were also employed to investigate the role of ROS and PERK/eIF-2α/ATF4 pathway in RANKL-induced autophagy and osteoclastogenesis. Results: RANKL markedly increased autophagy, while CQ treatment caused reduction of RANKL-induced autophagy and osteoclastogenesis. Consistent with the increased autophagy, the protein levels of Nox4 were significantly increased, and Nox4 was selectively localized within the endoplasmic reticulum (ER) after RANKL stimulation. DPI and shRNA efficiently decreased the protein level and (or) activity of Nox4 in the ER and inhibited RANKL-induced autophagy and osteoclastogenesis. Mechanistically, we found that Nox4 regulates RANKL-induced autophagy activation and osteoclastogenesis by stimulating the production of nonmitochondrial ROS. Additionally, Nox4-derived nonmitochondrial ROS dramatically activate PERK/eIF-2α/ATF4, which is a critical unfolded protein response (UPR)-related signaling pathway during ER stress. Blocking the activation of the PERK/eIF-2α/ATF4 signaling pathway either by Nox4 shRNA, ROS antioxidant or PERK inhibitor (GSK2606414) treatment significantly inhibited endoplasmic reticulum autophagy (ER-phagy) during RANKL-induced osteoclastogenesis. Conclusions: Our findings provide new insights into the processes of RANKL-induced osteoclastogenesis and will help the development of new therapeutic strategies for osteoclastogenesis-related diseases.

scholarly journals Degradation of a Short-lived Glycoprotein from the Lumen of the Endoplasmic Reticulum: The Role of N-linked Glycans and the Unfolded Protein Response

1999 ◽  
Vol 10 (12) ◽  
pp. 4059-4073 ◽  
Author(s):  
Maddalena de Virgilio ◽  
Claudia Kitzmüller ◽  
Eva Schwaiger ◽  
Michael Klein ◽  
Gert Kreibich ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Unfolded Protein Response ◽  
The Other ◽  
Slow Phase ◽  
Type I ◽  
Unfolded Protein ◽  
Other Hand ◽  
The Unfolded Protein Response ◽  
Protein Response

We are studying endoplasmic reticulum–associated degradation (ERAD) with the use of a truncated variant of the type I ER transmembrane glycoprotein ribophorin I (RI). The mutant protein, RI332, containing only the N-terminal 332 amino acids of the luminal domain of RI, has been shown to interact with calnexin and to be a substrate for the ubiquitin-proteasome pathway. When RI332 was expressed in HeLa cells, it was degraded with biphasic kinetics; an initial, slow phase of ∼45 min was followed by a second phase of threefold accelerated degradation. On the other hand, the kinetics of degradation of a form of RI332 in which the single used N-glycosylation consensus site had been removed (RI332-Thr) was monophasic and rapid, implying a role of the N-linked glycan in the first proteolytic phase. RI332degradation was enhanced when the binding of glycoproteins to calnexin was prevented. Moreover, the truncated glycoprotein interacted with calnexin preferentially during the first proteolytic phase, which strongly suggests that binding of RI332 to the lectin-like protein may result in the slow, initial phase of degradation. Additionally, mannose trimming appears to be required for efficient proteolysis of RI332. After treatment of cells with the inhibitor of N-glycosylation, tunicamycin, destruction of the truncated RI variants was severely inhibited; likewise, in cells preincubated with the calcium ionophore A23187, both RI332 and RI332-Thr were stabilized, despite the presence or absence of the N-linked glycan. On the other hand, both drugs are known to trigger the unfolded protein response (UPR), resulting in the induction of BiP and other ER-resident proteins. Indeed, only in drug-treated cells could an interaction between BiP and RI332 and RI332-Thr be detected. Induction of BiP was also evident after overexpression of murine Ire1, an ER transmembrane kinase known to play a central role in the UPR pathway; at the same time, stabilization of RI332 was observed. Together, these results suggest that binding of the substrate proteins to UPR-induced chaperones affects their half lives.

scholarly journals Multiple Myeloma-Derived Extracellular Vesicles Induce Osteoclastogenesis through the Activation of the XBP1/IRE1α Axis

Cancers ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2167 ◽  
Author(s):  
Lavinia Raimondi ◽  
Angela De Luca ◽  
Simona Fontana ◽  
Nicola Amodio ◽  
Viviana Costa ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Endoplasmic Reticulum ◽  
Extracellular Vesicle ◽  
Scientific Data ◽  
Daily Experience ◽  
Unfolded Protein ◽  
Xbp1 Mrna ◽  
Protein Response ◽  
Upr Pathway

Bone disease severely affects the quality of life of over 70% of multiple myeloma (MM) patients, which daily experience pain, pathological fractures, mobility issues and an increased mortality. Recent data have highlighted the crucial role of the endoplasmic reticulum-associated unfolded protein response (UPR) in malignant transformation and tumor progression; therefore, targeting of UPR-related molecules may open novel therapeutic avenues. Endoplasmic reticulum (ER) stress and UPR pathways are constitutively activated in MM cells, which are characterized by an increased protein turnover as a consequence of high production of immunoglobulins and high rates of protein synthesis. A great deal of scientific data also evidenced that a mild activation of UPR pathway can regulate cellular differentiation. Our previous studies revealed that MM cell-derived small extracellular vesicle (MM-EV) modulated osteoclasts (OCs) function and induced OCs differentiation. Here, we investigated the role of the UPR pathway, and in particular of the IRE1α/XBP1 axis, in osteoclastogenesis induced by MM-EVs. By proteomic analysis, we identified UPR signaling molecules as novel MM-EV cargo, prompting us to evaluate the effects of the MM-EVs on osteoclastogenesis through UPR pathway. MM-EVs administration in a murine macrophage cell line rapidly induced activation of IRE1α by phosphorylation in S724; accordingly, Xbp1 mRNA splicing was increased and the transcription of NFATc1, a master transcription factor for OCs differentiation, was activated. Some of these results were also validated using both human primary OC cultures and MM-EVs from MM patients. Notably, a chemical inhibitor of IRE1α (GSK2850163) counteracted MM-EV-triggered OC differentiation, hampering the terminal stages of OCs differentiation and reducing bone resorption.

scholarly journals Unfolded protein response in brain ischemia: A timely update

2016 ◽  
Vol 36 (12) ◽  
pp. 2044-2050 ◽  
Author(s):  
Wei Yang ◽  
Wulf Paschen
Keyword(s):  
Endoplasmic Reticulum ◽  
Endoplasmic Reticulum Stress ◽  
Unfolded Protein Response ◽  
Brain Ischemia ◽  
Unfolded Protein ◽  
Vital Functions ◽  
Advance Research ◽  
The Unfolded Protein Response ◽  
Protein Response

Folding and processing newly synthesized proteins are vital functions of the endoplasmic reticulum that are sensitive to a variety of stress conditions. The unfolded protein response is activated to restore endoplasmic reticulum function impaired by stress. While we know that brain ischemia impairs endoplasmic reticulum function, the role of unfolded protein response activation in post-ischemic recovery of neurologic function is only beginning to emerge. Here, we summarize what is known about endoplasmic reticulum stress and unfolded protein response in brain ischemia and discuss recent findings from myocardial ischemia studies that could help to advance research on endoplasmic reticulum stress and unfolded protein response in brain ischemia.

scholarly journals Regulation of the unfolded protein response by microRNAs

2013 ◽  
Vol 18 (4) ◽  
Author(s):  
Sylwia Bartoszewska ◽  
Kinga Kochan ◽  
Piotr Madanecki ◽  
Arkadiusz Piotrowski ◽  
Renata Ochocka ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Unfolded Protein Response ◽  
Potential Role ◽  
Recovery Phase ◽  
Cellular Mechanisms ◽  
Unfolded Protein ◽  
Critical Components ◽  
The Unfolded Protein Response ◽  
Protein Response

AbstractThe unfolded protein response (UPR) is an adaptive response to the stress that is caused by an accumulation of misfolded proteins in the lumen of the endoplasmic reticulum (ER). It is an important component of cellular homeostasis. During ER stress, the UPR increases the protein-folding capacity of the endoplasmic reticulum to relieve the stress. Failure to recover leads to apoptosis. Specific cellular mechanisms are required for the cellular recovery phase after UPR activation. Using bioinformatics tools, we identified a number of microRNAs that are predicted to decrease the mRNA expression levels for a number of critical components of the UPR. In this review, we discuss the potential role of microRNAs as key regulators of this pathway and describe how microRNAs may play an essential role in turning off the UPR after the stress has subsided.

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