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Molecular Cell Dec 2023N-glycans act as quality control tags by recruiting lectin chaperones to assist protein maturation in the endoplasmic reticulum. The location and composition of...
N-glycans act as quality control tags by recruiting lectin chaperones to assist protein maturation in the endoplasmic reticulum. The location and composition of N-glycans (glyco-code) are key to the chaperone-selection process. Serpins, a class of serine protease inhibitors, fold non-sequentially to achieve metastable active states. Here, the role of the glyco-code in assuring successful maturation and quality control of two human serpins, alpha-1 antitrypsin (AAT) and antithrombin III (ATIII), is described. We find that AAT, which has glycans near its N terminus, is assisted by early lectin chaperone binding. In contrast, ATIII, which has more C-terminal glycans, is initially helped by BiP and then later by lectin chaperones mediated by UGGT reglucosylation. UGGT action is increased for misfolding-prone disease variants, and these clients are preferentially glucosylated on their most C-terminal glycan. Our study illustrates how serpins utilize N-glycan presence, position, and composition to direct their proper folding, quality control, and trafficking.
Topics: Humans; Molecular Chaperones; Protein Folding; Lectins; Polysaccharides; Quality Control
PubMed: 38052210
DOI: 10.1016/j.molcel.2023.11.006 -
The FEBS Journal Dec 2023Functional networks in cells are created by physical, genetic, and regulatory interactions. Mapping them and annotating their functions by available methods remains a...
Functional networks in cells are created by physical, genetic, and regulatory interactions. Mapping them and annotating their functions by available methods remains a challenge. We use affinity purification mass spectrometry (AP-MS) coupled with SLiMFinder to discern such a network involving 26S proteasome non-ATPase regulatory subunit 9 (PSMD9), a chaperone of proteasome assembly. Approximately 20% of proteins within the PSMD9 interactome carry a short linear motif (SLiM) of the type 'EXKK'. The binding of purified PSMD9 with the peptide sequence ERKK, proteins heterogeneous nuclear ribonucleoproteins A2/B1 (hnRNPA2B1; containing ERKK), and peroxiredoxin-6 (PRDX6; containing EAKK) provided proof of principle for this motif-driven network. The EXKK motif in the peptide primarily interacts with the coiled-coil N domain of PSMD9, a unique interaction not reported for any coiled-coil domain. PSMD9 knockout (KO) HEK293 cells experience endoplasmic reticulum (ER) stress and respond by increasing the unfolded protein response (UPR) and reducing the formation of aggresomes and lipid droplets. Trans-expression of PSMD9 in the KO cells rescues lipid droplet formation. Overexpression of PSMD9 in HEK293 cells results in reduced UPR, and increased lipid droplet and aggresome formation. The outcome argues for the prominent role of PSMD9 in maintaining proteostasis. Probable mechanisms involve the binding of PSMD9 to binding immunoglobulin protein (BIP/GRP78; containing EDKK), an endoplasmic reticulum chaperone and key regulator of the UPR, and fatty acid synthase (FASN; containing ELKK), involved in fatty acid synthesis/lipid biogenesis. We propose that PSMD9 acts as a buffer in the cellular milieu by moderating the UPR and enhancing aggresome formation to reduce stress-induced proteotoxicity. Akin to waves created in ponds that perpetuate to a distance, perturbing the levels of PSMD9 would cause ripples down the networks, affecting final reactions in the pathway, one of which is altered proteostasis.
Topics: Humans; Proteasome Endopeptidase Complex; Proteostasis; HEK293 Cells; Molecular Chaperones; Unfolded Protein Response; Endoplasmic Reticulum Stress; Endoplasmic Reticulum Chaperone BiP; Carrier Proteins; Peptides
PubMed: 37665644
DOI: 10.1111/febs.16948 -
HSPB8 frameshift mutant aggregates weaken chaperone-assisted selective autophagy in neuromyopathies.Autophagy Aug 2023Chaperone-assisted selective autophagy (CASA) is a highly selective pathway for the disposal of misfolding and aggregating proteins. In muscle, CASA assures muscle...
Chaperone-assisted selective autophagy (CASA) is a highly selective pathway for the disposal of misfolding and aggregating proteins. In muscle, CASA assures muscle integrity by favoring the turnover of structural components damaged by mechanical strain. In neurons, CASA promotes the removal of aggregating substrates. A crucial player of CASA is HSPB8 (heat shock protein family B (small) member 8), which acts in a complex with HSPA, their cochaperone BAG3, and the E3 ubiquitin ligase STUB1. Recently, four novel frameshift (fs) gene mutations have been linked to neuromyopathies, and encode carboxy-terminally mutated HSPB8, sharing a common C-terminal extension. Here, we analyzed the biochemical and functional alterations associated with the HSPB8_fs mutant proteins. We demonstrated that HSPB8_fs mutants are highly insoluble and tend to form proteinaceous aggregates in the cytoplasm. Notably, all HSPB8 frameshift mutants retain their ability to interact with CASA members but sequester them into the HSPB8-positive aggregates together with two autophagy receptors SQSTM1/p62 and TAX1BP1. This copartitioning process negatively affects the CASA capability to remove its clients and causes a general failure in proteostasis response. Further analyses revealed that the aggregation of the HSPB8_fs mutants occurs independently of the other CASA members or from the autophagy receptors interaction, but it is an intrinsic feature of the mutated amino acid sequence. HSPB8_fs mutants aggregation alters the differentiation capacity of muscle cells and impairs sarcomere organization. Collectively, these results shed light on a potential pathogenic mechanism shared by the HSPB8_fs mutants described in neuromuscular diseases. : ACD: α-crystallin domain; ACTN: actinin alpha; BAG3: BAG cochaperone 3; C: carboxy; CASA: chaperone-assisted selective autophagy; CE: carboxy-terminal extension; CLEM: correlative light and electron microscopy; CMT2L: Charcot-Marie-Tooth type 2L; CTR: carboxy-terminal region; dHMNII: distal hereditary motor neuropathy type II; EV: empty vector; FRA: filter retardation assay; fs: frameshift; HSPA/HSP70: heat shock protein family A (Hsp70); HSPB1/Hsp27: heat shock protein family B (small) member 1; HSPB8/Hsp22: heat shock protein family B (small) member 8; HTT: huntingtin; KO: knockout; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MD: molecular dynamics; MTOC: microtubule organizing center; MYH: myosin heavy chain; MYOG: myogenin; NBR1: NBR1 autophagy cargo receptor; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; NSC34: Neuroblastoma X Spinal Cord 34; OPTN: optineurin; polyQ: polyglutamine; SQSTM1/p62: sequestosome 1; STUB1/CHIP: STIP1 homology and U-box containing protein 1; TARDBP/TDP-43: TAR DNA binding protein; TAX1BP1: Tax1 binding protein 1; TUBA: tubulin alpha; WT: wild-type.
Topics: Humans; Sequestosome-1 Protein; Autophagy; Heat-Shock Proteins; Neuromuscular Diseases; Charcot-Marie-Tooth Disease; Ubiquitin-Protein Ligases; Adaptor Proteins, Signal Transducing; Apoptosis Regulatory Proteins; Molecular Chaperones
PubMed: 36854646
DOI: 10.1080/15548627.2023.2179780 -
Biomolecules Dec 2023Alzheimer's disease (AD) is an extremely devastating neurodegenerative disease, and there is no cure for it. AD is specified as the misfolding and aggregation of... (Review)
Review
Alzheimer's disease (AD) is an extremely devastating neurodegenerative disease, and there is no cure for it. AD is specified as the misfolding and aggregation of amyloid-β protein (Aβ) and abnormalities in hyperphosphorylated tau protein. Current approaches to treat Alzheimer's disease have had some success in slowing down the disease's progression. However, attempts to find a cure have been largely unsuccessful, most likely due to the complexity associated with AD pathogenesis. Hence, a shift in focus to better understand the molecular mechanism of Aβ processing and to consider alternative options such as chaperone proteins seems promising. Chaperone proteins act as molecular caretakers to facilitate cellular homeostasis under standard conditions. Chaperone proteins like heat shock proteins (Hsps) serve a pivotal role in correctly folding amyloid peptides, inhibiting mitochondrial dysfunction, and peptide aggregation. For instance, Hsp90 plays a significant role in maintaining cellular homeostasis through its protein folding mechanisms. In this review, we analyze the most recent studies from 2020 to 2023 and provide updates on Aβ regulation by Hsp90, BRICHOS domain chaperone, and distinctive newly reported chaperones.
Topics: Humans; Alzheimer Disease; Heat-Shock Proteins; HSP90 Heat-Shock Proteins; Molecular Chaperones; Neurodegenerative Diseases; Amyloid beta-Peptides
PubMed: 38254616
DOI: 10.3390/biom14010016 -
International Journal of Biological... Dec 2023The aberrant accumulation of tau protein is implicated as a pathogenic factor in many neurodegenerative diseases. Tau seeding may underlie its predictable spread in...
The aberrant accumulation of tau protein is implicated as a pathogenic factor in many neurodegenerative diseases. Tau seeding may underlie its predictable spread in these diseases. Molecular chaperones can modulate tau pathology, but their effects have mainly been studied in isolation. This study employed a semi-high throughput assay to identify molecular chaperones influencing tau seeding using Tau RD P301S FRET Biosensor cells, which express a portion of tau containing the frontotemporal dementia-related P301S tau mutation fused to a FRET biosensor. Approximately fifty chaperones from five major families were screened using live cell imaging to monitor FRET-positive tau seeding. Among the tested chaperones, five exhibited significant effects on tau in the primary screen. Notably, three of these were from the DnaJ family. In subsequent studies, overexpression of DnaJA2, DnaJB1, and DnaJB6b resulted in significant reductions in tau levels. Knockdown experiments by shRNA revealed an inverse correlation between DnaJB1 and DnaJB6b with tau levels. DnaJB6b overexpression, specifically, reduced total tau levels in a cellular model with a pre-existing pool of tau, partially through enhanced proteasomal degradation. Further, DnaJB6b interacted with tau complexes. These findings highlight the potent chaperone activity within the DnaJ family, particularly DnaJB6b, towards tau.
Topics: Humans; tau Proteins; Frontotemporal Dementia; HSP40 Heat-Shock Proteins
PubMed: 37852393
DOI: 10.1016/j.ijbiomac.2023.127486 -
International Journal of Molecular... Feb 2024Alpha-B-crystallin, a member of the small heat shock family of proteins, has been implicated in a variety of cardiomyopathies and in normal cardiac homeostasis. It is... (Review)
Review
Alpha-B-crystallin, a member of the small heat shock family of proteins, has been implicated in a variety of cardiomyopathies and in normal cardiac homeostasis. It is known to function as a molecular chaperone, particularly for desmin, but also interacts with a wide variety of additional proteins. The molecular chaperone function is also enhanced by signal-dependent phosphorylation at specific residues under stress conditions. Naturally occurring mutations in , the gene that encodes alpha-B-crystallin, have been suggested to alter ionic intermolecular interactions that affect dimerization and chaperone function. These mutations have been associated with myofibrillar myopathy, restrictive cardiomyopathy, and hypertrophic cardiomyopathy and promote pathological hypertrophy through different mechanisms such as desmin aggregation, increased reductive stress, or activation of calcineurin-NFAT signaling. This review will discuss the known mechanisms by which alpha-B-crystallin functions in cardiac homeostasis and the pathogenesis of cardiomyopathies and provide insight into potential future areas of exploration.
Topics: Humans; Desmin; Cardiomyopathies; Mutation; Cardiomyopathy, Restrictive; Molecular Chaperones
PubMed: 38474073
DOI: 10.3390/ijms25052826 -
Cytokine & Growth Factor Reviews Feb 2024Hepatocellular carcinoma (HCC) is one of the most common gastrointestinal malignancies, characterized by insidious onset and high propensity for metastasis and... (Review)
Review
Hepatocellular carcinoma (HCC) is one of the most common gastrointestinal malignancies, characterized by insidious onset and high propensity for metastasis and recurrence. Apart from surgical resection, there are no effective curative methods for HCC in recent years, due to resistance to radiotherapy and chemotherapy. Heat shock proteins (HSP) play a crucial role in maintaining cellular homeostasis and normal organism development as molecular chaperones for intracellular proteins. Both basic research and clinical data have shown that HSPs are crucial participants in the HCC microenvironment, as well as the occurrence, development, metastasis, and resistance to radiotherapy and chemotherapy in various malignancies, particularly liver cancer. This review aims to discuss the molecular mechanisms and potential clinical value of HSPs in HCC, which may provide new insights for HSP-based therapeutic interventions for HCC.
Topics: Humans; Heat-Shock Proteins; Liver Neoplasms; Carcinoma, Hepatocellular; Molecular Chaperones; Tumor Microenvironment
PubMed: 38182465
DOI: 10.1016/j.cytogfr.2023.12.003 -
Cell Stress & Chaperones Sep 2023Chaperone proteins have crucial roles to play in all animal species and are involved in mediating both the folding of newly synthesized peptides into their mature... (Review)
Review
Chaperone proteins have crucial roles to play in all animal species and are involved in mediating both the folding of newly synthesized peptides into their mature conformation, the refolding of misfolded proteins, and the trafficking of proteins between subcellular compartments. These highly conserved proteins have particularly important roles to play in dealing with disruptions of the proteome as a result of environmental stress since abiotic factors, including temperature, pressure, oxygen, water availability, and pollutants can readily disrupt the conformation and/or function of all types of proteins, e.g., enzymes, transporters, and structural proteins. The current review provides an update on recent advances in understanding the roles and responses of chaperones in aiding animals to deal with environmental stress, offering new information on chaperone action in supporting survival strategies including torpor, hibernation, anaerobiosis, estivation, and cold/freeze tolerance among both vertebrate and invertebrate species.
Topics: Animals; Heat-Shock Proteins; Molecular Chaperones; Protein Folding; Cold Temperature; Temperature
PubMed: 36441380
DOI: 10.1007/s12192-022-01312-x -
Clinical and Translational Medicine Sep 2023Triple-negative breast cancer (TNBC), a heterogeneous subtype of breast cancer (BC), had poor prognosis. Endoplasmic reticulum (ER) stress was responsible for cellular...
BACKGROUND
Triple-negative breast cancer (TNBC), a heterogeneous subtype of breast cancer (BC), had poor prognosis. Endoplasmic reticulum (ER) stress was responsible for cellular processes and played a crucial role in the cell function. ER stress is a complex and dynamic process that can induce abnormal apoptosis and death. However, the underlying mechanism of ER stress involved in TNBC is not well defined.
METHODS
We identified ubiquitin-specific protease 19 (USP19) as a TNBC negative regulator for further investigation. The effects of USP19 on BC proliferation were assessed in vitro using proliferation test and cell-cycle assays, while the effects in vivo were examined using a mouse tumorigenicity model. Through in vitro flow cytometric analyses and in vivo TUNEL assays, cell apoptosis was assessed. Proteomics was used to examine the proteins that interact with USP19.
RESULTS
Multiple in vitro and in vivo tests showed that USP19 decreases TNBC cell growth while increasing apoptosis. Then, we demonstrated that USP19 interacts with deubiquitinates and subsequently stabilises family molecular chaperone regulator 6 (BAG6). BAG6 can boost B-cell lymphoma 2 (BCL2) ubiquitination and degradation, thereby raising ER calcium (Ca ) levels and causing ER stress. We also found that the N -methyladenosine (m A) "writer" methyltransferase-like 14 (METTL14) increased global m A modification.
CONCLUSIONS
Our study reveals that USP19 elevates the intracellular Ca concentration to alter ER stress via regulation of BAG6 and BCL2 stability and may be a viable therapeutic target for TNBC therapy.
Topics: Humans; Animals; Calcium; Triple Negative Breast Neoplasms; Endoplasmic Reticulum Stress; Disease Models, Animal; Deubiquitinating Enzymes; Proto-Oncogene Proteins c-bcl-2; Molecular Chaperones; Endopeptidases
PubMed: 37700495
DOI: 10.1002/ctm2.1398 -
Molecular and Cellular Biochemistry Oct 2023Chaperone-mediated autophagy (CMA) is the selective degradation process of intracellular components by lysosomes, which is required for the degradation of... (Review)
Review
Chaperone-mediated autophagy (CMA) is the selective degradation process of intracellular components by lysosomes, which is required for the degradation of aggregate-prone proteins and contributes to proteostasis maintenance. Proteostasis is essential for normal cell function and survival, and it is determined by the balance of protein synthesis and degradation. Because postmitotic neurons are highly susceptible to proteostasis disruption, CMA is vital for the nervous system. Since Parkinson's disease (PD) was first linked to CMA dysfunction, an increasing number of studies have shown that CMA loss, as seen during aging, occurs in the pathogenetic process of neurodegenerative diseases. Here, we review the molecular mechanisms of CMA, as well as the physiological function and regulation of this autophagy pathway. Following, we highlight its potential role in neurodegenerative diseases, and the latest advances and challenges in targeting CMA in therapy of neurodegenerative diseases.
Topics: Humans; Neurodegenerative Diseases; Chaperone-Mediated Autophagy; Molecular Chaperones; Autophagy; Parkinson Disease; Lysosomes
PubMed: 36695937
DOI: 10.1007/s11010-022-04640-9