Ubiquitin signals autophagic degradation of cytosolic proteins and peroxisomes.
Ontology highlight
ABSTRACT:
SUBMITTER: Kim PK
PROVIDER: S-EPMC2602605 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Peroxisome abundance is regulated by homeostasis between the peroxisomal biogenesis and degradation processes. Peroxin 16 (PEX16) is a peroxisomal protein involved in trafficking membrane proteins for de novo peroxisome biogenesis. The present study demonstrates that PEX16 also modulates peroxisome abundance through pexophagic degradation. PEX16 knockdown in human retinal pigment epithelial-1 cells decreased peroxisome abundance and function, represented by reductions in the expression of peroxisome membrane protein ABCD3 and the levels of cholesterol and plasmalogens, respectively. The activation of pexophagy under PEX16 knockdown was shown by (i) abrogated peroxisome loss under PEX16 knockdown in autophagy-deficient ATG5 knockout cell lines, and (ii) increased autophagy flux and co-localization of p62-an autophagy adaptor protein-with ABCD3 in the presence of the autophagy inhibitor chloroquine. However, the levels of cholesterol and plasmalogens did not recover despite the restoration of peroxisome abundance following chloroquine treatment. Thus, PEX16 is indispensable for maintaining peroxisome homeostasis by regulating not only the commonly known biogenesis pathway but also the autophagic degradation of peroxisomes.
| S-EPMC8348608 | biostudies-literature
Project description:Cellular protein quality control (PQC) systems selectively target misfolded or otherwise aberrant proteins for degradation by the ubiquitin-proteasome system (UPS). How cells discern abnormal from normal proteins remains incompletely understood, but involves in part the recognition between ubiquitin E3 ligases and degradation signals (degrons) that are exposed in misfolded proteins. PQC is compartmentalized in the cell, and a great deal has been learned in recent years about ER-associated degradation (ERAD) and nuclear quality control. In contrast, a comprehensive view of cytosolic quality control (CytoQC) has yet to emerge, and will benefit from the development of a well-defined set of model substrates. In this study, we generated an isogenic "degron library" in Saccharomyces cerevisiae consisting of short sequences appended to the C-terminus of a reporter protein, Ura3 About half of these degron-containing proteins are substrates of the integral membrane E3 ligase Doa10, which also plays a pivotal role in ERAD and some nuclear protein degradation. Notably, some of our degron fusion proteins exhibit dependence on the E3 ligase Ltn1/Rkr1 for degradation, apparently by a mechanism distinct from its known role in ribosomal quality control of translationally paused proteins. Ubr1 and San1, E3 ligases involved in the recognition of some misfolded CytoQC substrates, are largely dispensable for the degradation of our degron-containing proteins. Interestingly, the Hsp70/Hsp40 chaperone/cochaperones Ssa1,2 and Ydj1, are required for the degradation of all constructs tested. Taken together, the comprehensive degron library presented here provides an important resource of isogenic substrates for testing candidate PQC components and identifying new ones.
| S-EPMC4938640 | biostudies-literature
Project description:Damaged or redundant peroxisomes and their luminal cargoes are removed by pexophagy, a selective autophagy pathway. In yeasts, pexophagy depends mostly on the pexophagy receptors, such as Atg30 for Pichia pastoris and Atg36 for Saccharomyces cerevisiae, the autophagy scaffold proteins, Atg11 and Atg17, and the core autophagy machinery. In P. pastoris, the receptors for peroxisomal matrix proteins containing peroxisomal targeting signals (PTSs) include the PTS1 receptor, Pex5, and the PTS2 receptor and co-receptor, Pex7 and Pex20, respectively. These shuttling receptors are predominantly cytosolic and only partially peroxisomal. It remains unresolved as to whether, when and how the cytosolic pools of peroxisomal receptors, as well as the peroxisomal matrix proteins, are degraded under pexophagy conditions. These cytosolic pools exist both in normal and mutant cells impaired in peroxisome biogenesis. We report here that Pex5 and Pex7, but not Pex20, are degraded by an Atg30-independent, selective autophagy pathway. To enter this selective autophagy pathway, Pex7 required its major PTS2 cargo, Pot1. Similarly, the degradation of Pex5 was inhibited in cells missing abundant PTS1 cargoes, such as alcohol oxidases and Fox2 (hydratase-dehydrogenase-epimerase). Furthermore, in cells deficient in PTS receptors, the cytosolic pools of peroxisomal matrix proteins, such as Pot1 and Fox2, were also removed by Atg30-independent, selective autophagy, under pexophagy conditions. In summary, the cytosolic pools of PTS receptors and their cargoes are degraded via a pexophagy-independent, selective autophagy pathway under pexophagy conditions. These autophagy pathways likely protect cells from futile enzymatic reactions that could potentially cause the accumulation of toxic cytosolic products.Abbreviations: ATG: autophagy related; Cvt: cytoplasm to vacuole targeting; Fox2: hydratase-dehydrogenase-epimerase; PAGE: polyacrylamide gel electrophoresis; Pot1: thiolase; PMP: peroxisomal membrane protein; Pgk1: 3-phosphoglycerate kinase; PTS: peroxisomal targeting signal; RADAR: receptor accumulation and degradation in the absence of recycling; RING: really interesting new gene; SDS: sodium dodecyl sulphate; TCA, trichloroacetic acid; Ub: ubiquitin; UPS: ubiquitin-proteasome system Vid: vacuole import and degradation.
| S-EPMC6984484 | biostudies-literature
Project description:Protein quality control systems protect cells against the accumulation of toxic misfolded proteins by promoting their selective degradation. Malfunctions of quality control systems are linked to aging and neurodegenerative disease. Folding of polypeptides is facilitated by the association of 70 kDa Heat shock protein (Hsp70) molecular chaperones. If folding cannot be achieved, Hsp70 interacts with ubiquitylation enzymes that promote the proteasomal degradation of the misfolded protein. However, the factors that direct Hsp70 substrates toward the degradation machinery have remained unknown. Here, we identify Fes1, an Hsp70 nucleotide exchange factor of hitherto unclear physiological function, as a cytosolic triaging factor that promotes proteasomal degradation of misfolded proteins. Fes1 selectively interacts with misfolded proteins bound by Hsp70 and triggers their release from the chaperone. In the absence of Fes1, misfolded proteins fail to undergo polyubiquitylation, aggregate, and induce a strong heat shock response. Our findings reveal that Hsp70 direct proteins toward either folding or degradation by using distinct nucleotide exchange factors.
| S-EPMC3625341 | biostudies-literature
Project description:BackgroundCancer-associated cachexia and muscle wasting are considered key determinants of cancer-related death and reduction in the quality of life of cancer patients. A crucial link has been established between activin signaling and skeletal muscle atrophy-hypertrophy. We previously showed that activin-βC, a novel activin-A antagonist, is a tumor modulator that abolishes the cancer-associated cachexia in a mouse genetic model of gonadal tumorigenesis, in which the normal balance of inhibin/activin signalling is disrupted by a targeted mutation in the Inha gene (inhibin α-KO mouse). This study aimed to identify the molecular mechanism by which activin-βC increases survival and abolishes cancer-associated cachexia in α-KO mice. We hypothesized that overexpression of activin-βC modulates the cachexia phenotype by antagonizing the activin signaling pathway and repressing muscle wasting via the ubiquitin-proteasome and the autophagic-lysosomal degradation pathways.MethodsMale and female ActC++, α-KO, and α-KO/ActC++ mice and WT littermate controls were studied. Western blot analysis for the specific E3 ubiquitin ligases, atrogin-1 and MuRF1, markers of the autophagic-lysosomal pathway, Beclin-1, p62, and LC3A/B, effectors Smad-2, Smad-3 and myostatin was performed in the gastrocnemius of age-matched mice. Histopathology of the gastrocnemius and survival analysis were also conducted in animals from the same breeding cohort. Serum levels of activin-A, inflammatory cytokines, hormonal profile, and bone density were also assessed.ResultsIncreased levels of atrogin-1, MuRF-1, Beclin-1, p62, LC3A/B-I, Smad-2 and serum levels of activin-A were noted in the α-KO mice. These mice developed gonadal cancers followed by severe weight loss, and reduced survival. Overexpression of activin- βC antagonized the activin signaling cascade, attenuating the ubiquitin-proteasome and the autophagic-lysosomal degradation pathways, and reduced serum levels of activin-A. α-KO/ActC++ mice displayed a less aggressive cachectic phenotype, reduced tumor weight, and prolonged survival.ConclusionOur findings show for the first time a specific effect of activin-βC on muscle wasting and transcription factors involved in muscle protein degradation. The study indicates that activin-βC may be a novel therapy to abrogate cancer-associated weight loss and prolong survival.
| S-EPMC4670746 | biostudies-literature
Project description:The ubiquitin-proteasome pathway of protein degradation is one of the major mechanisms that are involved in the maintenance of the proper levels of cellular proteins. The regulation of proteasomal degradation thus ensures proper cell functions. The family of proteins containing ubiquitin-like (UbL) and ubiquitin-associated (UBA) domains has been implicated in proteasomal degradation. UbL-UBA domain containing proteins associate with substrates destined for degradation as well as with subunits of the proteasome, thus regulating the proper turnover of proteins.
| S-EPMC2725189 | biostudies-literature
Project description:Peroxisomes are organelles with crucial functions in oxidative metabolism. To correctly target to peroxisomes, proteins require specialized targeting signals. A mystery in the field is the sorting of proteins that carry a targeting signal for peroxisomes and as well as for other organelles, such as mitochondria or the endoplasmic reticulum (ER). Exploring several of these proteins in fungal model systems, we observed that they can act as tethers bridging organelles together to create contact sites. We show that in Saccharomyces cerevisiae this mode of tethering involves the peroxisome import machinery, the ER-mitochondria encounter structure (ERMES) at mitochondria and the guided entry of tail-anchored proteins (GET) pathway at the ER. Our findings introduce a previously unexplored concept of how dual affinity proteins can regulate organelle attachment and communication.
| S-EPMC10906886 | biostudies-literature
Project description:In eukaryotic cells an uncleavable ubiquitin moiety conjugated to the N-terminus of a protein signals the degradation of the fusion protein via the proteasome-dependent ubiquitin fusion degradation (UFD) pathway. In yeast the molecular mechanism of the UFD pathway has been well characterized. Recently the human E3 ubiquitin-protein ligase TRIP12 was connected with the UFD pathway, but little is otherwise known about this system in mammalian cells. In the present work, we utilized high-throughput imaging on cells transfected with a targeted siRNA library to identify components involved in degradation of the UFD substrate Ub(G76V)-YFP. The most significant hits from the screen were the E3 ubiquitin-protein ligase HUWE1, as well as PSMD7 and PSMD14 that encode proteasome subunits. Accordingly, knock down of HUWE1 led to an increase in the steady state level and a retarded degradation of the UFD substrate. Knock down of HUWE1 also led to a stabilization of the physiological UFD substrate UBB(+1). Precipitation experiments revealed that HUWE1 is associated with both the Ub(G76V)-YFP substrate and the 26S proteasome, indicating that it functions late in the UFD pathway. Double knock down of HUWE1 and TRIP12 resulted in an additive stabilization of the substrate, suggesting that HUWE1 and TRIP12 function in parallel during UFD. However, even when both HUWE1 and TRIP12 are downregulated, ubiquitylation of the UFD substrate was still apparent, revealing functional redundancy between HUWE1, TRIP12 and yet other ubiquitin-protein ligases.
| S-EPMC3507821 | biostudies-literature
Project description:A wide range of diseases are associated with the accumulation of cytosolic protein aggregates. The effects of these aggregates on various aspects of normal cellular protein homeostasis remain to be determined. Here we find that cytosolic aggregates, without necessarily disrupting proteasome function, can markedly delay the normally rapid degradation of nontranslocated secretory and membrane protein precursors. In the case of mammalian prion protein (PrP), the nontranslocated fraction is recruited into preexisting aggregates before its triage for degradation. This recruitment permits the growth and persistence of cytosolic PrP aggregates, explaining their apparent "self-conversion" seen in earlier studies of transient proteasome inhibition. For other proteins, the aggregate-mediated delay in precursor degradation led to aggregation and/or soluble residence in the cytosol, often causing aberrant cellular morphology. Remarkably, improving signal sequence efficiency mitigated these effects of aggregates. These observations identify a previously unappreciated consequence of cytosolic aggregates for nontranslocated secretory and membrane proteins, a minor but potentially disruptive population the rapid disposal of which is critical to maintaining cellular homeostasis.
| S-EPMC3093316 | biostudies-literature
Project description:An unbalanced karyotype, a condition known as aneuploidy, has a profound impact on cellular physiology and is a hallmark of cancer. Determining how aneuploidy affects cells is thus critical to understanding tumorigenesis. Here we show that aneuploidy interferes with the degradation of autophagosomes within lysosomes. Mis-folded proteins that accumulate in aneuploid cells due to aneuploidy-induced proteomic changes overwhelm the lysosome with cargo, leading to the observed lysosomal degradation defects. Importantly, aneuploid cells respond to lysosomal saturation. They activate a lysosomal stress pathway that specifically increases the expression of genes needed for autophagy-mediated protein degradation. Our results reveal lysosomal saturation as a universal feature of the aneuploid state that must be overcome during tumorigenesis. RPE-1 cells either untreated or treated with one of Reversine, Bafilomycin A1 or MG132, each condition was done in triplicate. D14-*_Control: untreated control D14-*_Rev: cells treated with 0.5uM Reversine for 24hrs and harvested 48hrs later D14-*_Baf: cells treated with 0.1uM BafA1 for 6hrs D14-*_Mg: cells treated with 1uM MG132 for 24 hrs
2015-10-26 | E-GEOD-60570 | biostudies-arrayexpress