Cell and Tissue Biology

, Volume 4, Issue 1, pp 36–53 | Cite as

Comparison of crude lysate pellets from isogenic strains of yeast with different prion composition: Identification of prion-associated proteins

  • O. V. Nevzglyadova
  • A. V. Artemov
  • A. G. Mittenberg
  • E. I. Kostyleva
  • E. V. Mikhailova
  • K. V. Solovyov
  • I. M. Kuznetsova
  • K. K. Turoverov
  • T. R. Soidla


A new approach involving the comparative analysis of proteins of crude cell lysate pellets from isogenic strains of Saccharomyces cerevisiae distinguished by their prion composition permitted us to identify a large group of prion-associated proteins in yeast cells. 35 proteins whose aggregation state depends on prion content have been identified by 2D-electrophoresis followed by the MALDI analysis of a recipient [psi ] strain and of [PSI +] cytoductant. Approximately half of these proteins belong to functional groups of chaperones and enzymes involved in glucose metabolism. Other proteins are involved in translation, stress response and protein degradation. The data obtained are compared with the results of other groups who used different approaches to detect proteins involved in prion aggregates.

Key words

amyloid prion heterokaryon cytoduction Saccharomyces cerevisiae 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Baci, L., Bertini, I., Boca, M., Girotto, S., Martinelli, M., Valentine, J.S., and Vieru, M., SOD1 and Amyotrophic Lateral Sclerosis: Mutations and Oligomerization. PLoS ONE, 2008, vol. 3, pp. e1677.CrossRefGoogle Scholar
  2. Bagriantsev, S.N., Gracheva, E.O., Richmond, J.E., and Liebman, S.W., Variant-Specific [PSI+] Infection Is Transmitted by Sup35 Polymers within [PSI +] Aggregates with Heterogeneous Protein Composition. Mol. Biol. Cell., 2008, vol. 19, pp. 2433–2443.CrossRefPubMedGoogle Scholar
  3. Buchan, J.R., Muhlrad, D., and Parker, R., P. Bodies Promote Stress Granule Assembly in Saccharomyces cerevisiae. J. Cell. Biol., 2008, vol. 183, pp. 441–455.CrossRefPubMedGoogle Scholar
  4. Cashikar, A.G., Duennwald, M., and Lindquist, S.L., A Chaperone Pathway in Protein Disaggregation: HSP26 Alters the Nature of Protein Aggregates to Facilitate Reactivation by HSP104. J. Biol. Chem., 2005, vol. 280, pp. 23869–23875.CrossRefPubMedGoogle Scholar
  5. Chernoff, Y.O., Cellular Control of Prion Formation and Propagation in Yeast, in Prions and Prion Diseases: Current Perspectives, Wymondham: Horizon Bioscience, 2004, pp. 257–303.Google Scholar
  6. Chernoff, Y.O., Identity Determinants of Infectious Proteins. Proc. Natl. Acad. Sci. USA, 2008, vol. 105, pp. 13191–13192.CrossRefPubMedGoogle Scholar
  7. Chernoff, Y.O., Lindquist, S.L., Ono, B., Inge-Vechtomov, S.G., and Liebman, S.W., Role of the Chaperone Protein hsp104 in Propagation of the Yeast Prion-Like Factor [PSI +]. Science, 1995, vol. 268, pp. 880–884.CrossRefPubMedGoogle Scholar
  8. Chernoff, Y.O., Newnam, G.P., Kumar, J., Allen, K., and Zink, A.D., Evidence of a Protein Mutator in Yeast: Role of the Hsp70-Related Chaperone Ssb in Formation, Stability, and Toxicity of the [PSI] Prion. Mol. Cell. Biol., 1999, vol. 19, pp. 8103–8112.PubMedGoogle Scholar
  9. Chernoff, Y.O., Uptain, S.M., and Lindquist, S.L., Analysis of Prion Factors in Yeast. Meth. Enzymol., 2002, vol. 351, pp. 499–537.CrossRefPubMedGoogle Scholar
  10. Chernova, T.A., Allen, K.D., Wesoloski, L.M., Shanks, J.R., Chernoff, Y.O., and Wilkinson, K.D., Pleiotropic Effects of Ubp6 Loss on Drug Sensitivities and Yeast Prion Are Due to Depletion of the Free Ubiquitin Pool. J. Biol. Chem., 2003, vol. 278, pp. 52102–52115.CrossRefPubMedGoogle Scholar
  11. Cox, B.S., Byrne, L., and Tuite, M.F., Prion Stability, in Protein-Based Inheritance, Austin, Texas: Landes Bioscience, 2007, pp. 56–72.Google Scholar
  12. Derkatch, I.L. and Liebman, S.W., Prion-Prion Interactions, in Protein-Based Inheritance, Austin, Texas: Landes Bioscience, 2007, pp. 39–55.Google Scholar
  13. Du, Z., Park, K.-W., Yu, H., Fan, Q., and Li, L., Newly Identified Prion Linked to the Chromatin-Remodeling Factor Swi1 in Saccharomyces cerevisiae, Nature Genetics, 2008, vol. 40, pp. 460–465.CrossRefGoogle Scholar
  14. Elam, J.S., Taylor, A.B., Strange, R., Antonyuk, S., Doucette, P.A., Rodriguez, J.A., Hasnain, S.S., Hayward, L.J., Valentine, J.S., Yeates, T.O., and Hart, P.J., Amyloid-Like Filaments and Water-Filled Nanotubes Formed by SOD1 Mutant Proteins Linked to Familial ALS. Nat. Struct. Biol., 2003, vol. 10, pp. 461–467.CrossRefPubMedGoogle Scholar
  15. Erjavec, N., Larsson, L., Grantham, J., and Nystrom, T., Accelerated Aging and Failure to Segregate Damaged Proteins in Sir2 Mutants Can Be Suppressed by Overproducing the Protein Aggregation-Remodeling Factor Hsp104p. Genes Dev., 2007, vol. 21, pp. 2410–2421.CrossRefPubMedGoogle Scholar
  16. Glabe, C.G., Statistical Classification of Toxic Amyloid Oligomers. J. Biol. Chem., 2008, vol. 283, pp. 29639–29643.CrossRefPubMedGoogle Scholar
  17. Griffith, J.S., Self-Replication and Scrapie. Nature, 1967, vol. 215, pp. 1943–1044.Google Scholar
  18. Harrison, L.B., Yu, Z., Stajich, J.E., Dietrich, F.S., and Harrison, P.M., Evolution of Budding Yeast Prion-Determinant Sequences across Diverse Fungi. J. Mol. Biol., 2007, vol. 368, pp. 273–282.CrossRefPubMedGoogle Scholar
  19. Herczenik, E. and Gebbink, M.F.B.G., Molecular and Cellular Aspects of Protein Misfolding and Disease. FASEB J., 2008, vol. 22, pp. 2115–2133.CrossRefPubMedGoogle Scholar
  20. Jones, G.W. and Masison, D.C.. Saccharomyces cerevisiae Hsp70 Mutations Affect [PSI+] Prion Propagation and Cell Growth Differently and Implicate Hsp40 and Tetratri-copeptide Repeat Cochaperones in Impairment of [PSI +]. Genetics, 2003, vol. 163, pp. 495–506.PubMedGoogle Scholar
  21. Jung, G., Jones, G., Wegrzyn, R.D., and Masison, D.C., A Role for Cytosolic Hsp70 in Yeast [PSI+] Prion Propagation and [PSI+] as a Cellular Stress. Genetics, 2000, vol. 156, pp. 559–570.PubMedGoogle Scholar
  22. Kryndushkin, D.S., Alexandrov, I.M., Ter-Avanesyan, M.D., and Kushnirov, V.V., Yeast [PSI+] Protein Aggregates Are Formed by Small Sup35 Polymers Fragmented by Hsp104. J. Biol. Chem., 2003, vol. 278, pp. 49636–49643.CrossRefPubMedGoogle Scholar
  23. Kushnirov, V.V., Alexandrov, I.M., Mitkevich, O.V., Shkundina, I.S., and Ter-Avanesyan, M.D., Purification and Analysis of Prion and Amyloid Aggregates. Methods, 2006, vol. 39, pp. 50–55.CrossRefPubMedGoogle Scholar
  24. Kushnirov, V.V., Vishnevskaya, A.B., Alexandrov, A.M., and Ter-Avanesyan, M.D., Prion and Nonprion Amyloids: A Comparison Inspired by the Yeast Sup35 Protein, in Protein-Based Inheritance, Austin, Texas: Landes Bioscience, 2007, pp. 73–82.Google Scholar
  25. Laemmli, U.K., Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4. Nature, 1970, vol. 227, pp. 680–685.CrossRefPubMedGoogle Scholar
  26. Lian, H.Y., Jiang, Y., Zhang, H., Jones, G.W., and Perrett, S., The Yeast Prion Protein Ure2: Structure, Functions, and Folding. Biochim. Biophys. Acta, 2006, vol. 1764, pp. 535–545.PubMedGoogle Scholar
  27. Lian, H.Y., Zhang, H., Zhang, Z.-R., Loovers, H.M., Jones, G.W., Rowling, P.J.E., Itzhaki, L.S., Zhou, J.-M., and Perrett, S., Hsp40 Interacts Directly with the Native State of the Yeast Prion Protein Ure2 and Inhibits Formation of Amyloid-Like Fibrils. J. Biol. Chem., 2007, vol. 282, pp. 11931–11940.CrossRefPubMedGoogle Scholar
  28. Nemecek, J., Nakayashiki, T., and Wickner, R.B., A Prion of Yeast Metacaspase Homolog (Mca1p) Detected by a Genetic Screen. Proc. Natl. Acad. Sci. USA, 2009, vol. 106, pp. 1892–1896.CrossRefPubMedGoogle Scholar
  29. Nevzglyadova, O.V., Artemov, A.V., Mittenberg, A.G., Mikhailova, E.V., Kuznetsova, I.M., Turoverov, K.K., and Soidla, T.R., Effect of the Red Pigment on Protein Amyloidization in Yeast. Tsitologiia, 2010a, vol. 51 (in press).Google Scholar
  30. Nevzglyadova, O.V., Artemov, A.V., Mittenberg, A.G., Mikhailova, E.V., Kuznetsova, I.M., Turoverov, K.K., and Soidla, T.R., Yeast Protein Aggregates, Containing Chaperones and Glucose Metabolism Enzymes, in Molecular Chaperones: Roles, Structures and Mechanisms, Hauppauge, New York: Nova Publ., 2010b.Google Scholar
  31. Nevzglyadova, O.V., Artyomov, A.V., Mikhailova, E.V., and Soidla, T.R., The Impact of Manipulations with Cytoplasmically Inherited Factors on Nuclear Transmission and Degradation in Yeast Heterokaryons. Curr. Genet., 2004, vol. 45, pp. 273–282.CrossRefPubMedGoogle Scholar
  32. Nevzglyadova, O.V., Kuznetsova, I.M., Artemov, A.V., Mikhailova, E.V., Turoverov, K.K., and Soidla, T.R., Estimating of Changes in the Amyloid and Prion Content of Yeast Cells. Tsitologiia, 2008, vol. 50 (1), pp. 40–48.Google Scholar
  33. Patel, B.K., Gavin-Smyth J., and Liebman, S.W., The Yeast Global Transcriptional Co-repressor Protein Cyc8 Can Propagate as a Prion. Nature Cell. Biol., 2009, vol 11, pp. 344–349.CrossRefPubMedGoogle Scholar
  34. Prion Biology and Diseases, Prusiner, S.B., Ed., 2nd ed., Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press, 2004.Google Scholar
  35. Protein-Based Inheritance, Chernoff, Y.O., Ed., Landes Bioscience: Austin, Texas, 2007.Google Scholar
  36. Rikhvanov, E.G., Romanova, N.V., and Chernoff, Y.O., Chaperone Effects on Prion and Nonprion Aggregates, in Protein-Based Inheritance, Austin, Texas: Landes Bioscience, 2007, pp. 83–92.Google Scholar
  37. Roberts, B.T. and Wickner, R.B., A Class of Prions That Propagate via Covalent Auto-activation. Genes Dev., 2003, vol. 17, pp. 2083–2087.CrossRefPubMedGoogle Scholar
  38. Satpute-Krishnan, P. and Serio, T.R., Prion Protein Remodeling Confers an Immediate Phenotype Switch. Nature, 2005, vol. 437, pp. 262–265.CrossRefPubMedGoogle Scholar
  39. Severin, S.E. and Solov’ev, G.A.. Praktikum po biokhimii (A Practical Course in Biochemistry), Moscow: Mosk. Gos. Univ., 1989.Google Scholar
  40. Sharma, D., Stanley, R.F., and Masison, D.C., Curing of Yeast [URE3] Prion by the Hsp40 Cochaperone Ydj1p Is Mediated by Hsp70. Genetics, 2009, vol. 181, pp. 129–137.CrossRefPubMedGoogle Scholar
  41. Sherman, F., Fink, G.R., and Hicks, J.B.. Laboratory Course Manual for Methods in Yeast Genetics, Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press, 1986.Google Scholar
  42. Sitia, R. and Molteni, S.N., Stress, Protein (Mis)folding, and Signaling: The Redox Connection. Sci. STKE, 2004, vol. 239, p. e27.Google Scholar
  43. Takahashi, A., Hara, H., Kurahashi, H., and Nakamura, Y., A Systematic Evaluation of the Function of the Protein-Remodeling Factor Hsp104 in [PSI+] Prion Propagation in S. cerevisiae by Comprehensive Chromosomal Mutations. Prion, 2007, vol. 1, pp. 69–77.PubMedGoogle Scholar
  44. Taneja, V., Maddelein, M.L., Talarek, N., Saupe, S.J., and Liebman, S.W., A Non-Q/N-Rich Prion Domain of a Foreign Prion, [Het-s], Can Propagate as a Prion in Yeast. Mol. Cell., 2007, vol. 27, pp. 67–77.CrossRefPubMedGoogle Scholar
  45. Towbin, H., Staehelin, T., and Gordon, J., Electrophoretic Transfer of Proteins from Polyacrylamide Gels to Nitrocellulose Sheets: Procedure and Some Applications. Proc. Natl. Acad. Sci. USA, 1979, vol. 76, pp. 4350–4354.CrossRefPubMedGoogle Scholar
  46. Tuite M.F. and Cox, B.S., The Genetic Control of the Formation and Propagation of the [PSI+] Prion in Yeast. Prion, 2007, vol. 1, pp. 101–109.PubMedGoogle Scholar
  47. Uversky, V.N., Oldfield, C.J., and Dunker, A.K., Intrinsically Disordered Proteins in Human Diseases: Introducing the D2 Concept. Annu. Rev. Biophys., 2008, vol. 37, pp. 215–246.CrossRefPubMedGoogle Scholar
  48. Uversky, V.N., Amyloidogenesis of Natively Unfolded Proteins. Curr. Alzheimer Res., 2008, vol. 5, pp. 260–287.CrossRefPubMedGoogle Scholar
  49. Wang, Y., Meriin, A.B., Costello, C.E., and Sherman, M.Y., Characterization of Proteins Associated with Polyglutamine Aggregates: A Novel Approach towards Isolation of Aggregates from Protein Conformation Disorders. Prion, 2007, vol. 1, pp. 128–135.PubMedGoogle Scholar
  50. Wang, Y., Meriin, A.B., Zaarur, N., Romanova, N.V., Chernoff, Y.O., Costello, C.E., and Sherman, M.Y., Abnormal Proteins Can Form Aggresome in Yeast: Aggresome-Targeting Signals and Components of the Machinery. FASEB J., 2008, vol. 23, pp. 451–463.CrossRefPubMedGoogle Scholar
  51. Watt, N.T., Taylor, D.R., Gillott, A., Thomas, D.A., Sumudhu, W., Perera, S., and Hooper, N.G., Reactive Oxygen Species-Mediated β-Cleavage of the Prion Protein in the Cellular Response to Oxidative Stress. J. Biol. Chem., 2005, vol. 280, pp. 35914–35921.CrossRefPubMedGoogle Scholar
  52. Wickner, R.B., [URE3] as an Altered URE2 Protein: Evidence for a Prion Analog in S. cerevisiae, Science, 1994, vol. 264, pp. 566–569.CrossRefPubMedGoogle Scholar
  53. Wickner, R.B., Edskes, H.K., and Shewmaker, F., How to Find a Prion: [URE3], [PSI +] and [beta]. Methods, 2006, vol. 39, pp. 3–8.CrossRefPubMedGoogle Scholar
  54. Wickner, R.B., Edskes, H.K., Shewmaker, F., Nakayashiki, T., Engel, A., McCann, L., and Kryndushkin, D., Yeast Prions: Evolution of the Prion Concept. Prion, 2007, vol. 1, pp. 94–100.PubMedGoogle Scholar
  55. Wickner, R.B., Shewmaker, F., Kryndushkin, D., and Edskes, H.K., Protein Inheritance (Prions) Based on Parallel In-register β-Sheet Amyloid Structures. BioEssays, 2008, vol. 30, pp. 955–964.CrossRefPubMedGoogle Scholar
  56. Zakharov, I.A. and Yarovoy, B., Cytoduction as a New Tool in Studying the Cytoplasmic Heredity in Yeast. Mol. Cell. Biochem., 1977, vol. 14, pp. 15–18.CrossRefPubMedGoogle Scholar
  57. Zhou, R.Y., Li, X.L., Li, L.H., Li, X.Y., and Feng, F.J., Evolution and Differentiation of the Prion Protein Gene (PRNP) among Species. J. Heredity, 2008, vol. 99, pp. 647–652.CrossRefGoogle Scholar

Copyright information

© Pleiades Publishing, Ltd. 2010

Authors and Affiliations

  • O. V. Nevzglyadova
    • 1
  • A. V. Artemov
    • 1
  • A. G. Mittenberg
    • 1
  • E. I. Kostyleva
    • 1
  • E. V. Mikhailova
    • 1
  • K. V. Solovyov
    • 2
  • I. M. Kuznetsova
    • 1
  • K. K. Turoverov
    • 1
  • T. R. Soidla
    • 1
  1. 1.Institute of CytologyRussian Academy of SciencesSt. PetersburgRussia
  2. 2.Institute of Experimental MedicineRussian Academy of Medical SciencesSt. PetersburgRussia

Personalised recommendations