Cell Stress and Chaperones

, Volume 17, Issue 1, pp 57–66

Reactivation of protein aggregates by mortalin and Tid1—the human mitochondrial Hsp70 chaperone system

  • Ohad Iosefson
  • Shelly Sharon
  • Pierre Goloubinoff
  • Abdussalam Azem
Original Paper

Abstract

The mitochondrial 70-kDa heat shock protein (mtHsp70), also known in humans as mortalin, is a central component of the mitochondrial protein import motor and plays a key role in the folding of matrix-localized mitochondrial proteins. MtHsp70 is assisted by a member of the 40-kDa heat shock protein co-chaperone family named Tid1 and a nucleotide exchange factor. Whereas, yeast mtHsp70 has been extensively studied in the context of protein import in the mitochondria, and the bacterial 70-kDa heat shock protein was recently shown to act as an ATP-fuelled unfolding enzyme capable of detoxifying stably misfolded polypeptides into harmless natively refolded proteins, little is known about the molecular functions of the human mortalin in protein homeostasis. Here, we developed novel and efficient purification protocols for mortalin and the two spliced versions of Tid1, Tid1-S, and Tid1-L and showed that mortalin can mediate the in vitro ATP-dependent reactivation of stable-preformed heat-denatured model aggregates, with the assistance of Mge1 and either Tid1-L or Tid1-S co-chaperones or yeast Mdj1. Thus, in addition of being a central component of the protein import machinery, human mortalin together with Tid1, may serve as a protein disaggregating machine which, for lack of Hsp100/ClpB disaggregating co-chaperones, may carry alone the scavenging of toxic protein aggregates in stressed, diseased, or aging human mitochondria.

Keywords

Mitochondrial Hsp70 Tid1 Hep1 Ssc1 Disaggregation 

Supplementary material

12192_2011_285_MOESM1_ESM.doc (842 kb)
Supplementary Fig. 1Sequence alignment of human mortalin with yeast mtHsp70s (Ssc1, Ssq1, and Ecm10) and E. coli DnaK. (DOC 842 kb)
12192_2011_285_MOESM2_ESM.doc (530 kb)
Supplementary Fig. 2Sequence alignment of human Tid1-L and Tid1-S, yeast mtHsp40 (Mdj1). and E coli DnaJ. (DOC 530 kb)
12192_2011_285_MOESM3_ESM.doc (1.2 mb)
Supplementary Fig. 3Purification steps of mortalin. (DOC 1,255 kb)
12192_2011_285_MOESM4_ESM.doc (1.4 mb)
Supplementary Fig. 4Purification steps of Tid1-L. (DOC 1,388 kb)
12192_2011_285_MOESM5_ESM.doc (52 kb)
Supplementary Fig. 5The ATPase activity of mortalin is not stimulated by Tid1-L H121Q, under single-turnover conditions. (DOC 52 kb)
12192_2011_285_MOESM6_ESM.doc (56 kb)
Supplementary Fig. 6Time-dependent reactivation of stable G6PDH aggregates by mortalin and mortalin-V482F. (DOC 56 kb)
12192_2011_285_MOESM7_ESM.doc (36 kb)
Supplementary Table 1Sequence of primers (DOC 36 kb)

References

  1. Aldridge JE, Horibe T, Hoogenraad NJ (2007) Discovery of genes activated by the mitochondrial unfolded protein response (mtUPR) and cognate promoter elements. PLoS One 2(9):e874. doi:10.1371/journal.pone.0000874 PubMedCrossRefGoogle Scholar
  2. Baumann F, Milisav I, Neupert W, Herrmann JM (2000) Ecm10, a novel hsp70 homolog in the mitochondrial matrix of the yeast Saccharomyces cerevisiae. FEBS Lett 487(2):307–312PubMedCrossRefGoogle Scholar
  3. Bhattacharyya T, Karnezis AN, Murphy SP, Hoang T, Freeman BC, Phillips B, Morimoto RI (1995) Cloning and subcellular localization of human mitochondrial hsp70. J Biol Chem 270(4):1705–1710PubMedCrossRefGoogle Scholar
  4. Blamowska M, Sichting M, Mapa K, Mokranjac D, Neupert W, Hell K (2010) ATPase domain and interdomain linker play a key role in aggregation of mitochondrial Hsp70 chaperone Ssc1. J Biol Chem 285(7):4423–4431. doi:10.1074/jbc.M109.061697 PubMedCrossRefGoogle Scholar
  5. Broadley SA, Hartl FU (2009) The role of molecular chaperones in human misfolding diseases. FEBS Lett 583(16):2647–2653. doi:10.1016/j.febslet.2009.04.029 PubMedCrossRefGoogle Scholar
  6. Burbulla LF, Schelling C, Kato H, Rapaport D, Woitalla D, Schiesling C, Schulte C, Sharma M, Illig T, Bauer P, Jung S, Nordheim A, Schols L, Riess O, Kruger R (2010) Dissecting the role of the mitochondrial chaperone mortalin in Parkinson’s disease: functional impact of disease-related variants on mitochondrial homeostasis. Hum Mol Genet. doi:10.1093/hmg/ddq370
  7. D’Silva P, Liu Q, Walter W, Craig EA (2004) Regulated interactions of mtHsp70 with Tim44 at the translocon in the mitochondrial inner membrane. Nat Struct Mol Biol 11(11):1084–1091. doi:10.1038/nsmb846 PubMedCrossRefGoogle Scholar
  8. Daugaard M, Rohde M, Jaattela M (2007) The heat shock protein 70 family: Highly homologous proteins with overlapping and distinct functions. FEBS Lett 581(19):3702–3710. doi:10.1016/j.febslet.2007.05.039 PubMedCrossRefGoogle Scholar
  9. De Castro IP, Martins LM, Tufi R (2010) Mitochondrial quality control and neurological disease: an emerging connection. Expert Rev Mol Med 12:e12. doi:10.1017/S1462399410001456 PubMedCrossRefGoogle Scholar
  10. De Los Rios P, Ben-Zvi A, Slutsky O, Azem A, Goloubinoff P (2006) Hsp70 chaperones accelerate protein translocation and the unfolding of stable protein aggregates by entropic pulling. Proc Natl Acad Sci USA 103(16):6166–6171. doi:10.1073/pnas.0510496103 CrossRefGoogle Scholar
  11. De Mena L, Coto E, Sanchez-Ferrero E, Ribacoba R, Guisasola LM, Salvador C, Blazquez M, Alvarez V (2009) Mutational screening of the mortalin gene (HSPA9) in Parkinson’s disease. J Neural Transm 116(10):1289–1293. doi:10.1007/s00702-009-0273-2 PubMedCrossRefGoogle Scholar
  12. Deloche O, Liberek K, Zylicz M, Georgopoulos C (1997) Purification and biochemical properties of Saccharomyces cerevisiae Mdj1p, the mitochondrial DnaJ homologue. J Biol Chem 272(45):28539–28544PubMedCrossRefGoogle Scholar
  13. Deocaris CC, Widodo N, Ishii T, Kaul SC, Wadhwa R (2007) Functional significance of minor structural and expression changes in stress chaperone mortalin. Ann N Y Acad Sci 1119:165–175. doi:10.1196/annals.1404.007 PubMedCrossRefGoogle Scholar
  14. Deocaris CC, Kaul SC, Wadhwa R (2008) From proliferative to neurological role of an hsp70 stress chaperone, mortalin. Biogerontology 9(6):391–403. doi:10.1007/s10522-008-9174-2 PubMedCrossRefGoogle Scholar
  15. Diamant S, Ben-Zvi AP, Bukau B, Goloubinoff P (2000) Size-dependent disaggregation of stable protein aggregates by the DnaK chaperone machinery. J Biol Chem 275(28):21107–21113. doi:10.1074/jbc.M001293200 PubMedCrossRefGoogle Scholar
  16. Dutkiewicz R, Schilke B, Knieszner H, Walter W, Craig EA, Marszalek J (2003) Ssq1, a mitochondrial Hsp70 involved in iron-sulfur (Fe/S) center biogenesis. Similarities to and differences from its bacterial counterpart. J Biol Chem 278(32):29719–29727. doi:10.1074/jbc.M303527200 PubMedCrossRefGoogle Scholar
  17. Genevaux P, Georgopoulos C, Kelley WL (2007) The Hsp70 chaperone machines of Escherichia coli: a paradigm for the repartition of chaperone functions. Mol Microbiol 66(4):840–857. doi:10.1111/j.1365-2958.2007.05961.x PubMedCrossRefGoogle Scholar
  18. Glover JR, Lindquist S (1998) Hsp104, Hsp70, and Hsp40: a novel chaperone system that rescues previously aggregated proteins. Cell 94(1):73–82PubMedCrossRefGoogle Scholar
  19. Goloubinoff P, De Los Rios P (2007) The mechanism of Hsp70 chaperones: (entropic) pulling the models together. Trends Biochem Sci 32(8):372–380. doi:10.1016/j.tibs.2007.06.008 PubMedCrossRefGoogle Scholar
  20. Goloubinoff P, Mogk A, Zvi AP, Tomoyasu T, Bukau B (1999) Sequential mechanism of solubilization and refolding of stable protein aggregates by a bichaperone network. Proc Natl Acad Sci USA 96(24):13732–13737PubMedCrossRefGoogle Scholar
  21. Goswami AV, Chittoor B, D’Silva P (2010) Understanding the functional interplay between mammalian mitochondrial Hsp70 chaperone machine components. J Biol Chem 285(25):19472–19482. doi:10.1074/jbc.M110.105957 PubMedCrossRefGoogle Scholar
  22. Hansen JE, Gafni A (1993) Thermal switching between enhanced and arrested reactivation of bacterial glucose-6-phosphate dehydrogenase assisted by GroEL in the absence of ATP. J Biol Chem 268(29):21632–21636PubMedGoogle Scholar
  23. Hinault MP, Farina-Henriquez-Cuendet A, Mattoo RU, Mensi M, Dietler G, Lashuel HA, Goloubinoff P (2010) Stable {alpha}-synuclein oligomers strongly inhibit chaperone activity of the HSP70 system by weak interactions with J-domain co-chaperones. J Biol Chem. doi:10.1074/jbc.M110.127753
  24. Horst M, Oppliger W, Rospert S, Schonfeld HJ, Schatz G, Azem A (1997) Sequential action of two hsp70 complexes during protein import into mitochondria. EMBO J 16(8):1842–1849. doi:10.1093/emboj/16.8.1842 PubMedCrossRefGoogle Scholar
  25. Iosefson O, Azem A (2010) Reconstitution of the mitochondrial Hsp70 (mortalin)-p53 interaction using purified proteins—identification of additional interacting regions. FEBS Lett. doi:10.1016/j.febslet.2010.02.019
  26. Kaul SC, Deocaris CC, Wadhwa R (2007) Three faces of mortalin: a housekeeper, guardian and killer. Exp Gerontol 42(4):263–274. doi:10.1016/j.exger.2006.10.020 PubMedCrossRefGoogle Scholar
  27. Koren J 3rd, Jinwal UK, Lee DC, Jones JR, Shults CL, Johnson AG, Anderson LJ, Dickey CA (2009) Chaperone signalling complexes in Alzheimer’s disease. J Cell Mol Med 13(4):619–630PubMedCrossRefGoogle Scholar
  28. Laufen T, Mayer MP, Beisel C, Klostermeier D, Mogk A, Reinstein J, Bukau B (1999) Mechanism of regulation of hsp70 chaperones by DnaJ cochaperones. Proc Natl Acad Sci USA 96(10):5452–5457PubMedCrossRefGoogle Scholar
  29. Liu Q, Krzewska J, Liberek K, Craig EA (2001) Mitochondrial Hsp70 Ssc1: role in protein folding. J Biol Chem 276(9):6112–6118. doi:10.1074/jbc.M009519200 PubMedCrossRefGoogle Scholar
  30. Lu B, Garrido N, Spelbrink JN, Suzuki CK (2006) Tid1 isoforms are mitochondrial DnaJ-like chaperones with unique carboxyl termini that determine cytosolic fate. J Biol Chem 281(19):13150–13158. doi:10.1074/jbc.M509179200 PubMedCrossRefGoogle Scholar
  31. Mapa K, Sikor M, Kudryavtsev V, Waegemann K, Kalinin S, Seidel CA, Neupert W, Lamb DC, Mokranjac D (2010) The conformational dynamics of the mitochondrial Hsp70 chaperone. Mol Cell 38(1):89–100. doi:10.1016/j.molcel.2010.03.010 PubMedCrossRefGoogle Scholar
  32. Mitra A, Shevde LA, Samant RS (2009) Multi-faceted role of HSP40 in cancer. Clin Exp Metastasis 26(6):559–567. doi:10.1007/s10585-009-9255-x PubMedCrossRefGoogle Scholar
  33. Mizzen LA, Chang C, Garrels JI, Welch WJ (1989) Identification, characterization, and purification of two mammalian stress proteins present in mitochondria, grp 75, a member of the hsp 70 family and hsp 58, a homolog of the bacterial groEL protein. J Biol Chem 264(34):20664–20675PubMedGoogle Scholar
  34. Mogk A, Tomoyasu T, Goloubinoff P, Rudiger S, Roder D, Langen H, Bukau B (1999) Identification of thermolabile Escherichia coli proteins: prevention and reversion of aggregation by DnaK and ClpB. EMBO J 18(24):6934–6949. doi:10.1093/emboj/18.24.6934 PubMedCrossRefGoogle Scholar
  35. Neupert W, Herrmann JM (2007) Translocation of proteins into mitochondria. Annu Rev Biochem 76:723–749. doi:10.1146/annurev.biochem.76.052705.163409 PubMedCrossRefGoogle Scholar
  36. Pedersen CB, Bross P, Winter VS, Corydon TJ, Bolund L, Bartlett K, Vockley J, Gregersen N (2003) Misfolding, degradation, and aggregation of variant proteins. The molecular pathogenesis of short chain acyl-CoA dehydrogenase (SCAD) deficiency. J Biol Chem 278(48):47449–47458. doi:10.1074/jbc.M309514200 PubMedCrossRefGoogle Scholar
  37. Rowley N, Prip-Buus C, Westermann B, Brown C, Schwarz E, Barrell B, Neupert W (1994) Mdj1p, a novel chaperone of the DnaJ family, is involved in mitochondrial biogenesis and protein folding. Cell 77(2):249–259PubMedCrossRefGoogle Scholar
  38. Schapira AH (1998) Mitochondrial dysfunction in neurodegenerative disorders. Biochim Biophys Acta 1366(1–2):225–233PubMedGoogle Scholar
  39. Sharma SK, Christen P, Goloubinoff P (2009) Disaggregating chaperones: an unfolding story. Curr Protein Pept Sci 10(5):432–446PubMedCrossRefGoogle Scholar
  40. Sharma SK, De Los Rios P, Christen P, Lustig A, Goloubinoff P (2010) The kinetic parameters and energy cost of the Hsp70 chaperone as a polypeptide unfoldase. Nat Chem Biol. doi:10.1038/nchembio.455
  41. Shi M, Jin J, Wang Y, Beyer RP, Kitsou E, Albin RL, Gearing M, Pan C, Zhang J (2008) Mortalin: a protein associated with progression of Parkinson disease? J Neuropathol Exp Neurol 67(2):117–124. doi:10.1097/nen.0b013e318163354a PubMedCrossRefGoogle Scholar
  42. Sichting M, Mokranjac D, Azem A, Neupert W, Hell K (2005) Maintenance of structure and function of mitochondrial Hsp70 chaperones requires the chaperone Hep1. EMBO J 24(5):1046–1056. doi:10.1038/sj.emboj.7600580 PubMedCrossRefGoogle Scholar
  43. Swain JF, Dinler G, Sivendran R, Montgomery DL, Stotz M, Gierasch LM (2007) Hsp70 chaperone ligands control domain association via an allosteric mechanism mediated by the interdomain linker. Mol Cell 26(1):27–39. doi:10.1016/j.molcel.2007.02.020 PubMedCrossRefGoogle Scholar
  44. Syken J, De-Medina T, Munger K (1999) TID1, a human homolog of the Drosophila tumor suppressor l(2)tid, encodes two mitochondrial modulators of apoptosis with opposing functions. Proc Natl Acad Sci USA 96(15):8499–8504PubMedCrossRefGoogle Scholar
  45. Szabo A, Langer T, Schroder H, Flanagan J, Bukau B, Hartl FU (1994) The ATP hydrolysis-dependent reaction cycle of the Escherichia coli Hsp70 system DnaK, DnaJ, and GrpE. Proc Natl Acad Sci USA 91(22):10345–10349PubMedCrossRefGoogle Scholar
  46. Vijayvergiya C, Beal MF, Buck J, Manfredi G (2005) Mutant superoxide dismutase 1 forms aggregates in the brain mitochondrial matrix of amyotrophic lateral sclerosis mice. J Neurosci 25(10):2463–2470. doi:10.1523/JNEUROSCI.4385-04.2005 PubMedCrossRefGoogle Scholar
  47. von Janowsky B, Major T, Knapp K, Voos W (2006) The disaggregation activity of the mitochondrial ClpB homolog Hsp78 maintains Hsp70 function during heat stress. J Mol Biol 357(3):793–807. doi:10.1016/j.jmb.2006.01.008 CrossRefGoogle Scholar
  48. Wadhwa R, Kaul SC, Ikawa Y, Sugimoto Y (1993) Identification of a novel member of mouse hsp70 family. Its association with cellular mortal phenotype. J Biol Chem 268(9):6615–6621PubMedGoogle Scholar
  49. Weibezahn J, Schlieker C, Tessarz P, Mogk A, Bukau B (2005) Novel insights into the mechanism of chaperone-assisted protein disaggregation. Biol Chem 386(8):739–744. doi:10.1515/BC.2005.086 PubMedCrossRefGoogle Scholar
  50. Xie W, Wan OW, Chung KK (2010) New insights into the role of mitochondrial dysfunction and protein aggregation in Parkinson’s disease. Biochim Biophys Acta 1802(11):935–941. doi:10.1016/j.bbadis.2010.07.014 PubMedGoogle Scholar
  51. Zhai P, Stanworth C, Liu S, Silberg JJ (2008) The human escort protein Hep binds to the ATPase domain of mitochondrial hsp70 and regulates ATP hydrolysis. J Biol Chem 283(38):26098–26106. doi:10.1074/jbc.M803475200 PubMedCrossRefGoogle Scholar
  52. Zhao Q, Wang J, Levichkin IV, Stasinopoulos S, Ryan MT, Hoogenraad NJ (2002) A mitochondrial specific stress response in mammalian cells. EMBO J 21(17):4411–4419PubMedCrossRefGoogle Scholar

Copyright information

© Cell Stress Society International 2011

Authors and Affiliations

  • Ohad Iosefson
    • 1
  • Shelly Sharon
    • 1
  • Pierre Goloubinoff
    • 2
  • Abdussalam Azem
    • 1
  1. 1.Department of Biochemistry and Molecular Biology, George S. Wise Faculty of Life SciencesTel Aviv UniversityTel AvivIsrael
  2. 2.Département de Biologie Moléculaire VégétaleUniversité de LausanneLausanneSwitzerland

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