Proteasome Inhibition in Wild-Type Yeast Saccharomyces Cerevisiae Cells
2007; Future Science Ltd; Volume: 42; Issue: 2 Linguagem: Inglês
10.2144/000112389
ISSN1940-9818
AutoresChang Liu, Jennifer Apodaca, Laura E. Davis, Hai Rao,
Tópico(s)Glycosylation and Glycoproteins Research
ResumoBioTechniquesVol. 42, No. 2 BenchmarksOpen AccessProteasome inhibition in wild-type yeast Saccharomyces cerevisiae cellsChang Liu, Jennifer Apodaca, Laura E. Davis & Hai RaoChang LiuThe University of Texas Health Science Center, San Antonio, TX, USA, Jennifer Apodaca*Address correspondence to: Jennifer Apodaca, Institute of Biotechnology, The University of Texas Health Science Center, 15355 Lambda Dr., San Antonio, TX 78245, USA. e-mail: E-mail Address: apodaca@uthscsa.eduThe University of Texas Health Science Center, San Antonio, TX, USA, Laura E. DavisThe University of Texas Health Science Center, San Antonio, TX, USA & Hai RaoThe University of Texas Health Science Center, San Antonio, TX, USAPublished Online:16 May 2018https://doi.org/10.2144/000112389AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinkedInReddit The lysosome and 26S proteasome represent the two major proteolytic machines in eukaryotic cells (1). While the lysosome deals mainly with nonse-lective proteolysis, the 26S proteasome handles the majority of regulated proteolysis. The 26S proteasome is a multisubunit protease that degrades the substrate into small peptides in an ATP-dependent manner (2,3). The prote-asome has three peptidase activities, including (i) chymotrypsin-like, (ii) trypsin-like, and (iii) peptidylglutamyl-peptide hydrolase activities (2).To demonstrate that a protein is a substrate of the proteasome in vivo, the stability of the protein is often examined and compared in the presence or absence of proteasome inhibitors (e.g., MG132), short peptide aldehydes that block active sites of the proteasome (4). Use of the budding yeast Saccharomyces cerevisiae as a model system has been instrumental in uncovering mechanistic attributes and the physiologic functions of the prote-asome. However, the use of proteasome inhibitors in wild-type S. cerevisiae cells is hampered by the impermeability of the cell wall or membrane (5). Therefore, mutant yeast strains (e.g., erg6Δ, pdr5Δ) with increased drug permeability or reduced drug efflux are required for experiments using proteasome inhibitors (5,6). A caveat to this approach is that mutation in ERG6 or PDR5 may directly or indirectly affect some cellular processes (e.g., increased import of sodium) (7,8) and protein stability. Furthermore, in some cases, the ERG6 or PDR5 genes must be deleted in another mutant background, a technically cumbersome step, to establish the involvement of the 26S proteasome in a particular process (e.g., transcription or telomere maintenance) (9,10). Recently, a method was developed involving brefeldin A, an antifungal agent often used to study protein trafficking from the endoplasmic reticulum (ER) to the Golgi apparatus; this method allowed the efficient uptake of brefeldin A in wild-type yeast cells (11). The key elements of this strategy are the use of L-proline instead of ammonium sulfate as the sole nitrogen source in the growth medium and the addition of a small amount of sodium dodecyl sulfate (SDS; 0.003%). These treatments likely lead to the transient opening of the cell wall/membrane, as yeast cells become permeable to brefeldin A and the dye crystal violet.We have adapted this simple method for inhibiting the proteasome in wild-type S. cerevisiae cells. Here, we demonstrate that the degradation of distinct proteasomal substrates can be blocked via this approach. Four proteasomal substrates tested are cytosolic proteins UVV76-β-galactosidase (β-gal) and Deg1-β-gal, and two misfolded ER membrane proteins, Ubc6 and Hmg2, that are ubiquitylated by different ubiquitin-protein ligases (12,13). These substrates have been routinely employed to study proteasome -mediated degradation.We used cycloheximide to terminate protein synthesis and followed the fate of these substrates in the presence or absence of the proteasome inhibitor MG132. Specifically, yeast cells expressing Deg1-β-gal, myc-tagged Hmg2, or Ha-tagged Ubc6 were grown at 30°C in a synthetic medium (0.17% yeast nitrogenous base without ammonium sulfate) supplemented with 0.1% proline, appropriate amino acids, and 2% glucose as the carbon source. The culture grown overnight was reinoculated into 30 mL fresh media with 0.003% SDS (electrophoresis grade; Fisher Scientific, Fair Lawn, NJ, USA) at A600 0.5. The cells were grown for an additional 3 h at 30°C. Then, cells were added with 75 µM MG132 (Biomol, Plymouth Meeting, PA, USA) or the control buffer dimethyl sulfoxide (DMSO). After a 30-min incubation, 100 µg/mL cycloheximide were added to yeast cells to stop protein synthesis and start the chase. Samples were withdrawn at the indicated time points and harvested by centrifugation at 2520x g for 5 min. Cells were resuspended in lysis buffer (50 mM HEPES, pH 7.5, 5 mM EDTA, 150 mM NaCl, 1% Triton® X-100, protease inhibitor mix) and lysed by glass beads. Protein concentration was determined by the Bradford assay. Equal amounts of proteins were resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) (Figure 1, A and C). To detect myc-tagged Hmg2 (Figure 1B), immunoprecipitations were performed by mixing extracts with the beads coated with myc antibody (9E10) for 2 h at 4°C. Gels were transferred to a polyvinylidene difluoride (PVDF) membrane. Immunoblots were probed with monoclonal antibody (1:4000 dilution) against β-gal (Sigma-Aldrich, St. Louis, MO, USA), myc-, or Ha-epitope (Covance Research Products, Berkley, CA, USA), then the goat anti-mouse horseradish peroxidase (HRP) conjugate, and were developed using ECL® reagents (GE Healthcare, Piscataway, NJ, USA) as previously described (14). The stable protein Rpt5 was employed as the loading control to ensure equal amounts of extracts were used (Figure 1, A-C). Consistent with previous reports, Deg1-β-gal, Hmg2, and Ubc6 are degraded in wild-type cells (Figure 1, A-C). Addition of proteasome inhibitor MG132 significantly compromised the degradation of these proteins (Figure 1, A-C), suggesting that this method efficiently impairs proteasome activity.Figure 1. Effects of the proteasome inhibitor MG132 on the degradation of short-lived proteins in wild-type yeast cells.Wild-type yeast cells expressing (A) Deg1-β-galactosidase (β-gal), (B) myc-Hmg2, or (C) HA-Ubc6 were treated with or without the proteasome inhibitor MG132 (75 µM). Time points were taken after expression shutoff. Proteins were extracted from these cells and separated on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) (A and C). Extracts containing myc-Hmg2 were immunoprecipitated with beads coupled to myc antibody to enrich myc-Hmg2; immunoprecipitates were resolved by SDS-PAGE (B). After the electrophoresis, gels were transferred to a polyvinylidene difluoride (PVDF) membrane and then probed with appropriate antibodies. To ensure that equal amounts of extracts were used in the experiments, extracts were similarly separated on SDS-PAGE, and the immu-noblots were probed with the stable protein Rpt5 as the loading control (A–C). (D) SDS and proline are required for efficient inhibition of ubiquitin fusion degradation (UFD) substrate turnover by MG132. Wild-type cells expressing UVV76-β-gal were grown in media with or without SDS and proline. MG132 (40 µM) was used in these experiments. UVV76-β-gal stability was determined as described above and detected by anti-β-gal antibody. An arrow indicates UVV76-β-gal. Ubiquitylated UVV76-β-gal species are seen above the arrow. (E) Dose response to MG132 treatment. Wild-type yeast cells expressing UVV76-β-gal were grown in various concentrations of MG132 in the presence of SDS and proline. Levels of β-gal activity were determined as previously described (15). The experiments were done more than three times, and the average values with standard deviation are shown.To determine the amount of MG132 required for efficient proteasome inhibition, we employed the model substrate UVV76-β-gal, which is degraded by the ubiquitin fusion degradation (UFD) pathway. We used the lacZ assay to gauge the effects of MG132 on the intracellular concentration of the UFD substrate (15). We found that potent inhibition of UVV76-β-gal degradation can be achieved with approximately 20–40 µM MG132 (Figure 1E). Furthermore, we also used cycloheximide chase to compare the degradation of the UFD substrate in the presence or absence of the SDS and proline treatment (Figure 1D). The addition of SDS and proline, which did not significantly alter substrate degradation on their own (data not shown), is essential for the MG132-induced stabilization of UFD substrates (Figure 1D).In this report, we have demonstrated that the treatment of proline and SDS allows MG132 to effectively inhibit the proteasome in wild-type cells. In addition, we found that another proteasome inhibitor MG262 (4) can also block proteasome-mediated degradation of UFD substrates (data not shown). Though the addition of SDS and proline in the growth media caused approximately 26% reduced viability (11), this method eliminates the need for mutation in ERG6 or PDR5, which may alter normal cellular events with undesired effects (7,8). The strategy described could be used directly to demonstrate the role of the proteasome in a specific pathway without the cumbersome need to generate double mutants (9). Moreover, as it is often challenging to characterize proteins that are rapidly degraded by the prote-asome, this strategy would help detect protein-protein interactions or ubiqui-tylated forms of these substrates.AcknowledgmentsC.L. and J.A. contributed equally to this work. We are grateful to Drs. M. Hochstrasser and R. Hampton for plas-mids. We thank D. Sharp, B. Christy, M. Gaczynska, and I. Kim for support. H.R. was supported by grants from the American Cancer Society (RSG-05-158-01-TBE), the Barshop Center for Aging Studies (P30AG13319-10), and The University of Texas Health Science Center (UTHSC) Institutional Grants.Competing Interests StatementThe authors declare no competing interests.References1. Pickart, C.M. and R.E. Cohen. 2004. Proteasomes and their kin: proteases in the machine age. Nat. Rev. Mol. Cell Biol. 5:177–187.Crossref, Medline, CAS, Google Scholar2. DeMartino, G.N. and C.A. Slaughter. 1999. The proteasome, a novel protease regulated by multiple mechanisms. J. Biol. Chem. 274:22123–22126.Crossref, Medline, CAS, Google Scholar3. Goldberg, A.L., S.J. Elledge, and J.W. Harper. 2001. The cellular chamber of doom. Sci. Am. 284:68–73 .Crossref, Medline, CAS, Google Scholar4. Gaczynska, M. and P.A. Osmulski. 2005. Small-molecule inhibitors of proteasome activity. Methods Mol. Biol. 301:3–22.Medline, CAS, Google Scholar5. Lee, D.H. and A.L. Goldberg. 1996. Selective inhibitors of the proteasome-depen-dent and vacuolar pathways of protein degradation in Saccharomyces cerevisiae. J. Biol. Chem. 271:27280–27284.Crossref, Medline, CAS, Google Scholar6. Fleming, J.A., E.S. Lightcap, S. Sadis, V. Thoroddsen, C.E. Bulawa, and R.K. Blackman. 2002. Complementary whole-genome technologies reveal the cellular response to proteasome inhibition by PS-341. Proc. Natl. Acad. Sci. USA 99:1461–1466.Crossref, Medline, CAS, Google Scholar7. Gaber, R.F., D.M. Copple, B.K. Kennedy, M. Vidal, and M. Bard. 1989. The yeast gene ERG6 is required for normal membrane function but is not essential for biosynthesis of the cell-cycle-sparking sterol. Mol. Cell. Biol. 9:3447–3456.Crossref, Medline, CAS, Google Scholar8. Welihinda, A.A., A.D. Beavis, and R.J. Trumbly. 1994. Mutations in LIS1 (ERG6) gene confer increased sodium and lithium uptake in Saccharomyces cerevisiae. Biochim. Biophys. Acta 1193:107–117.Crossref, Medline, CAS, Google Scholar9. Lipford, J.R., G.T. Smith, Y. Chi, and R.J. Deshaies. 2005. A putative stimulatory role for activator turnover in gene expression. Nature 438:113–116.Crossref, Medline, CAS, Google Scholar10. Osterhage, J.L., J.M. Talley, and K.L. Friedman. 2006. Proteasome-dependent degradation of Est1p regulates the cell cycle-restricted assembly of telomerase in Saccharomyces cerevisiae. Nat. Struct. Mol. Biol. 13:720–728.Crossref, Medline, CAS, Google Scholar11. Pannunzio, V.G., H.I. Burgos, M. Alonso, J.R. Mattoon, E.H. Ramos, and C.A. Stella. 2004. A simple chemical method for rendering wild-type yeast permeable to Brefeldin A that does not require the presence of an erg6 mutation. J. Biomed. Biotechnol. 2004:150–155.Crossref, Medline, Google Scholar12. Gardner, R.G., A.G. Shearer, and R.Y. Hampton. 2001. In vivo action of the HRD ubiquitin ligase complex: mechanisms of en-doplasmic reticulum quality control and sterol regulation. Mol. Cell. Biol. 21:4276–4291.Crossref, Medline, CAS, Google Scholar13. Ravid, T., S.G. Kreft, and M. Hochstrasser. 2006. Membrane and soluble substrates of the Doa10 ubiquitin ligase are degraded by distinct pathways. EMBO J. 25:533–543.Crossref, Medline, CAS, Google Scholar14. Kim, I., J. Ahn, C. Liu, K. Tanabe, J. Apodaca, T. Suzuki, and H. Rao. 2006. The Png1-Rad23 complex regulates glycoprotein turnover. J. Cell Biol. 172:211–219.Crossref, Medline, CAS, Google Scholar15. Kim, I., K. Mi, and H. Rao. 2004. Multiple interactions of Rad23 suggest a mechanism for ubiquitylated substrate delivery important in proteolysis. Mol. Biol. Cell 15:3357–3365.Crossref, Medline, CAS, Google ScholarFiguresReferencesRelatedDetailsCited BySumoylation is Largely Dispensable for Normal Growth but Facilitates Heat Tolerance in Yeast31 January 2023 | Molecular and Cellular Biology, Vol. 43, No. 1Processing of Fluorescent Proteins May Prevent Detection of Prion Particles in [PSI+] Cells22 November 2022 | Biology, Vol. 11, No. 12Redox-sensitive E2 Rad6 controls cellular response to oxidative stress via K63-linked ubiquitination of ribosomesCell Reports, Vol. 39, No. 8Developing systems in yeast to address Alzheimer's diseaseChemical rescue of mutant proteins in living Saccharomyces cerevisiae cells by naturally occurring small molecules17 July 2021 | G3 Genes|Genomes|Genetics, Vol. 11, No. 9Membrane dynamics and protein targets of lipid droplet microautophagy during ER stress-induced proteostasis in the budding yeast, Saccharomyces cerevisiae6 October 2020 | Autophagy, Vol. 17, No. 9Regulation of Cell Death Induced by Acetic Acid in Yeasts24 June 2021 | Frontiers in Cell and Developmental Biology, Vol. 9An Hsp90 co-chaperone links protein folding and degradation and is part of a conserved protein quality controlCell Reports, Vol. 35, No. 13Analysis of Protein Stability by Synthesis ShutoffBIO-PROTOCOL, Vol. 11, No. 22Homologous recombination and Mus81 promote replication completion in response to replication fork blockage17 May 2020 | EMBO reports, Vol. 21, No. 7The Cdc48-20S proteasome degrades a class of endogenous proteins in a ubiquitin-independent mannerBiochemical and Biophysical Research Communications, Vol. 523, No. 4A AAA ATPase Cdc48 with a cofactor Ubx2 facilitates ubiquitylation of a mitochondrial fusion-promoting factor Fzo1 for proteasomal degradation5 December 2019 | The Journal of Biochemistry, Vol. 167, No. 3Dual role of a GTPase conformational switch for membrane fusion by mitofusin ubiquitylation19 December 2019 | Life Science Alliance, Vol. 3, No. 1Sumoylation regulates the stability and nuclease activity of Saccharomyces cerevisiae Dna28 May 2019 | Communications Biology, Vol. 2, No. 1Local translation of yeast ERG4 mRNA at the endoplasmic reticulum requires the brefeldin A resistance protein Bfr127 August 2019 | RNA, Vol. 25, No. 12Polyubiquitin Chains Linked by Lysine Residue 48 (K48) Selectively Target Oxidized Proteins In VivoAntioxidants & Redox Signaling, Vol. 31, No. 15The Proteasome Lid Triggers COP9 Signalosome Activity during the Transition of Saccharomyces cerevisiae Cells into Quiescence4 September 2019 | Biomolecules, Vol. 9, No. 9A Flp-SUMO hybrid recombinase reveals multi-layered copy number control of a selfish DNA element through post-translational modification26 June 2019 | PLOS Genetics, Vol. 15, No. 6Nuclear proteasomal degradation of Saccharomyces cerevisiae inorganic pyrophosphatase Ipp1p, a nucleocytoplasmic protein whose stability depends on its subcellular localizationBiochimica et Biophysica Acta (BBA) - Molecular Cell Research, Vol. 1866, No. 6Heterologous Hsp90 promotes phenotypic diversity through network evolution15 November 2018 | PLOS Biology, Vol. 16, No. 11CLIP and cohibin separate rDNA from nucleolar proteins destined for degradation by nucleophagy29 June 2018 | Journal of Cell Biology, Vol. 217, No. 8Bre1 mediates the ubiquitination of histone H2B by regulating Lge1 stability24 April 2018 | FEBS Letters, Vol. 592, No. 9Cdc48 regulates a deubiquitylase cascade critical for mitochondrial fusion8 January 2018 | eLife, Vol. 7A Method to Monitor Protein Turnover by Flow Cytometry and to Screen for Factors that Control Degradation by Fluorescence-Activated Cell Sorting22 September 2018Investigation of an optimal cell lysis method for the study of the zinc metalloproteome of Histoplasma capsulatum12 August 2017 | Analytical and Bioanalytical Chemistry, Vol. 409, No. 26An ubiquitin-dependent balance between mitofusin turnover and fatty acids desaturation regulates mitochondrial fusion13 June 2017 | Nature Communications, Vol. 8, No. 1To CURe or not to CURe? Differential effects of the chaperone sorting factor Cur1 on yeast prions are mediated by the chaperone Sis19 May 2017 | Molecular Microbiology, Vol. 105, No. 2Histone degradation in response to DNA damage enhances chromatin dynamics and recombination rates9 January 2017 | Nature Structural & Molecular Biology, Vol. 24, No. 2Chromatin Association of Gcn4 Is Limited by Post-translational Modifications Triggered by its DNA-Binding in Saccharomyces cerevisiae1 December 2016 | Genetics, Vol. 204, No. 4Interaction of Gcn4 with target gene chromatin is modulated by proteasome functionMolecular Biology of the Cell, Vol. 27, No. 17Autophagic Turnover of Inactive 26S Proteasomes in Yeast Is Directed by the Ubiquitin Receptor Cue5 and the Hsp42 ChaperoneCell Reports, Vol. 16, No. 6Ubiquilin/Dsk2 promotes inclusion body formation and vacuole (lysosome)-mediated disposal of mutated huntingtinMolecular Biology of the Cell, Vol. 27, No. 13Degradation Signals for Ubiquitin-Proteasome Dependent Cytosolic Protein Quality Control (CytoQC) in Yeast1 July 2016 | G3 Genes|Genomes|Genetics, Vol. 6, No. 7C-Terminal Tyrosine Residue Modifications Modulate the Protective Phosphorylation of Serine 129 of α-Synuclein in a Yeast Model of Parkinson's Disease24 June 2016 | PLOS Genetics, Vol. 12, No. 6The Rqc2/Tae2 subunit of the ribosome-associated quality control (RQC) complex marks ribosome-stalled nascent polypeptide chains for aggregation4 March 2016 | eLife, Vol. 5Studying Protein Ubiquitylation in Yeast10 September 2016Wss1 metalloprotease partners with Cdc48/Doa1 in processing genotoxic SUMO conjugates8 September 2015 | eLife, Vol. 4Compromising the 19S proteasome complex protects cells from reduced flux through the proteasome1 September 2015 | eLife, Vol. 4Sch9 regulates intracellular protein ubiquitination by controlling stress responsesRedox Biology, Vol. 5Not4‐dependent translational repression is important for cellular protein homeostasis in yeast13 May 2015 | The EMBO Journal, Vol. 34, No. 14Pollen S-locus F-box proteins of Petunia involved in S-RNase-based self-incompatibility are themselves subject to ubiquitin-mediated degradation31 May 2015 | The Plant Journal, Vol. 83, No. 2Cdc48-independent proteasomal degradation coincides with a reduced need for ubiquitylation5 January 2015 | Scientific Reports, Vol. 5, No. 1The 26S Proteasome Degrades the Soluble but Not the Fibrillar Form of the Yeast Prion Ure2p In Vitro26 June 2015 | PLOS ONE, Vol. 10, No. 6Cysteine-specific ubiquitination protects the peroxisomal import receptor Pex5p against proteasomal degradation22 June 2015 | Bioscience Reports, Vol. 35, No. 3Sumoylation controls the timing of Tup1-mediated transcriptional deactivation13 March 2015 | Nature Communications, Vol. 6, No. 1Inhibiting K63 Polyubiquitination Abolishes No-Go Type Stalled Translation Surveillance in Saccharomyces cerevisiae24 April 2015 | PLOS Genetics, Vol. 11, No. 4Rad25 Protein Is Targeted for Degradation by the Ubc4-Ufd4 PathwayJournal of Biological Chemistry, Vol. 290, No. 13Interplay between Sumoylation and Phosphorylation for Protection against α-Synuclein InclusionsJournal of Biological Chemistry, Vol. 289, No. 45Unraveling the Biology of a Fungal Meningitis Pathogen Using Chemical GeneticsCell, Vol. 159, No. 5Phosphorylation by Casein Kinase 2 Facilitates Psh1 Protein-assisted Degradation of Cse4 ProteinJournal of Biological Chemistry, Vol. 289, No. 42Stress-dependent Proteolytic Processing of the Actin Assembly Protein Lsb1 Modulates a Yeast PrionJournal of Biological Chemistry, Vol. 289, No. 40Interactions of the natural product kendomycin and the 20S proteasomeJournal of Molecular Biology, Vol. 426, No. 18Timely Activation of Budding Yeast APCCdh1 Involves Degradation of Its Inhibitor, Acm1, by an Unconventional Proteolytic Mechanism29 July 2014 | PLoS ONE, Vol. 9, No. 7Non-repair Pathways for Minimizing Protein Isoaspartyl Damage in the Yeast Saccharomyces cerevisiaeJournal of Biological Chemistry, Vol. 289, No. 24Structural and Functional Profiling of the Lateral Gate of the Sec61 TransloconJournal of Biological Chemistry, Vol. 289, No. 22A role for the proteasome in the turnover of Sup35p and in [ PSI+ ] prion propagation17 March 2014 | Molecular Microbiology, Vol. 92, No. 3Ybp1 and Gpx3 Signaling in Candida albicans Govern Hydrogen Peroxide-Induced Oxidation of the Cap1 Transcription Factor and Macrophage EscapeAntioxidants & Redox Signaling, Vol. 19, No. 18TORC1 Inhibits GSK3-Mediated Elo2 Phosphorylation to Regulate Very Long Chain Fatty Acid Synthesis and AutophagyCell Reports, Vol. 5, No. 4The C-terminal Residues of Saccharomyces cerevisiae Mec1 Are Required for Its Localization, Stability, and Function1 October 2013 | G3 Genes|Genomes|Genetics, Vol. 3, No. 10Cisplatin-induced cell death in Saccharomyces cerevisiae is programmed and rescued by proteasome inhibitionDNA Repair, Vol. 12, No. 6The Ubiquitin-Proteasome System Regulates Mitochondrial Intermembrane Space ProteinsMolecular and Cellular Biology, Vol. 33, No. 11Two Deubiquitylases Act on Mitofusin and Regulate Mitochondrial Fusion along Independent PathwaysMolecular Cell, Vol. 49, No. 3Defective in Mitotic Arrest 1 (Dma1) Ubiquitin Ligase Controls G1 Cyclin DegradationJournal of Biological Chemistry, Vol. 288, No. 7The Ubiquitin–Proteasome System of Saccharomyces cerevisiae1 October 2012 | Genetics, Vol. 192, No. 2Saccharomyces cerevisiae Apn1 mutation affecting stable protein expression mimics catalytic activity impairment: Implications for assessing DNA repair capacity in humansDNA Repair, Vol. 11, No. 9Solubility-Promoting Function of Hsp90 Contributes to Client Maturation and Robust Cell GrowthEukaryotic Cell, Vol. 11, No. 8The Yeast Ubr1 Ubiquitin Ligase Participates in a Prominent Pathway That Targets Cytosolic Thermosensitive Mutants for Degradation1 May 2012 | G3 Genes|Genomes|Genetics, Vol. 2, No. 5The Ubiquitin Ligase Ubr11 Is Essential for Oligopeptide Utilization in the Fission Yeast Schizosaccharomyces pombeEukaryotic Cell, Vol. 11, No. 3Sumoylation of transcription factor Gcn4 facilitates its Srb10-mediated clearance from promoters in yeast16 February 2012 | Genes & Development, Vol. 26, No. 4Letter to the Editor8 December 2011 | Yeast, Vol. 29, No. 2C-terminal UBA domains protect ubiquitin receptors by preventing initiation of protein degradation8 February 2011 | Nature Communications, Vol. 2, No. 1The Cdc48 ATPase modulates the interaction between two proteolytic factors Ufd2 and Rad231 August 2011 | Proceedings of the National Academy of Sciences, Vol. 108, No. 33Ubp15p, a Ubiquitin Hydrolase Associated with the Peroxisomal Export MachineryJournal of Biological Chemistry, Vol. 286, No. 32Budding yeast Dma1 and Dma2 participate in regulation of Swe1 levels and localizationMolecular Biology of the Cell, Vol. 22, No. 13Cdc48/p97 Mediates UV-Dependent Turnover of RNA Pol IIMolecular Cell, Vol. 41, No. 1Seipin Is a Discrete Homooligomer18 November 2010 | Biochemistry, Vol. 49, No. 50Combined chemical and genetic approach to inhibit proteolysis by the proteasome2 November 2010 | Yeast, Vol. 27, No. 11Cks1, Cdk1, and the 19S Proteasome Collaborate To Regulate Gene Induction-Dependent Nucleosome Eviction in Yeast20 March 2023 | Molecular and Cellular Biology, Vol. 30, No. 22Degradation of the Saccharomyces cerevisiae Mating-Type Regulator α1: Genetic Dissection of Cis -determinants and Trans -acting Pathways1 June 2010 | Genetics, Vol. 185, No. 2Mutant p62/SQSTM1 UBA domains linked to Paget's disease of bone differ in their abilities to function as stabilization signals15 March 2010 | FEBS Letters, Vol. 584, No. 8Ubiquitin Chain Elongation Enzyme Ufd2 Regulates a Subset of Doa10 SubstratesJournal of Biological Chemistry, Vol. 285, No. 14Npr2, Yeast Homolog of the Human Tumor Suppressor NPRL2 , Is a Target of Grr1 Required for Adaptation to Growth on Diverse Nitrogen SourcesEukaryotic Cell, Vol. 9, No. 4A genome-wide synthetic dosage lethality screen reveals multiple pathways that require the functioning of ubiquitin-binding proteins Rad23 and Dsk212 November 2009 | BMC Biology, Vol. 7, No. 1The Nedd4-Type Rsp5p Ubiquitin Ligase Inhibits Tombusvirus Replication by Regulating Degradation of the p92 Replication Protein and Decreasing the Activity of the Tombusvirus ReplicaseJournal of Virology, Vol. 83, No. 22TORC1 controls degradation of the transcription factor Stp1, a key effector of the SPS amino-acid-sensing pathway in Saccharomyces cerevisiaeJournal of Cell Science, Vol. 122, No. 12Polyubiquitination of the demethylase Jhd2 controls histone methylation and gene expression3 April 2009 | Genes & Development, Vol. 23, No. 8Nucleus-Specific and Cell Cycle-Regulated Degradation of Mitogen-Activated Protein Kinase Scaffold Protein Ste5 Contributes to the Control of Signaling CompetenceMolecular and Cellular Biology, Vol. 29, No. 2Minimal length requirement for proteasomal degradation of ubiquitin‐dependent substrates16 September 2008 | The FASEB Journal, Vol. 23, No. 1 Vol. 42, No. 2 Follow us on social media for the latest updates Metrics History Received 25 August 2006 Accepted 30 November 2006 Published online 16 May 2018 Published in print February 2007 Information© 2007 Author(s)AcknowledgmentsC.L. and J.A. contributed equally to this work. We are grateful to Drs. M. Hochstrasser and R. Hampton for plas-mids. We thank D. Sharp, B. Christy, M. Gaczynska, and I. Kim for support. H.R. was supported by grants from the American Cancer Society (RSG-05-158-01-TBE), the Barshop Center for Aging Studies (P30AG13319-10), and The University of Texas Health Science Center (UTHSC) Institutional Grants.Competing Interests StatementThe authors declare no competing interests.PDF download
Referência(s)