**6. Acknowledgements**

We acknowledge financial support provided by the Spanish Ministry of Health according to the 'Plan Nacional de I+D+I 2008-2011', Instituto Nacional Carlos III (ISCIII, project CP10/00527 to C.R.), and cofunding by FEDER funds., and the PAIDI Program from the Andalusian Government (CTS-677). A.L.G. holds a FPU Fellowship from the Spanish Ministry of Science (MICINN). C.W.B. acknowledges funding from the EU FP7 (MC-IEF 236721). The authors are grateful to Christopher M. Dobson (University of Cambridge, UK) and John Christodoulou (University College London, UK) for helpful and highly stimulating discussion of results.

The Hsp70 Chaperone System in Parkinson's Disease 237

Burbulla, L. F., et al. (2010). Dissecting the role of the mitochondrial chaperone mortalin in

Ciechanover, A.&P. Brundin (2003). The ubiquitin proteasome system in neurodegenerative diseases: sometimes the chicken, sometimes the egg. *Neuron*, 40, (2), pp. (427-46). Connell, P., et al. (2001). The co-chaperone CHIP regulates protein triage decisions mediated

Conway, K. A., et al. (1998). Accelerated in vitro fibril formation by a mutant alphasynuclein linked to early-onset Parkinson disease. *Nat Med*, 4, (11), pp. (1318-20). Conway, K. A., et al. (2000). Acceleration of oligomerization, not fibrillization, is a shared

Cook, C.&L. Petrucelli (2009). A critical evaluation of the ubiquitin-proteasome system in

Cooper, A. A., et al. (2006). Alpha-synuclein blocks ER-Golgi traffic and Rab1 rescues

Cudkowicz, M. E., et al. (2008). Arimoclomol at dosages up to 300 mg/day is well tolerated and safe in amyotrophic lateral sclerosis. *Muscle Nerve*, 38, (1), pp. (837-44). Cuervo, A. M., et al. (2004). Impaired degradation of mutant alpha-synuclein by chaperone-

Chandra, S., et al. (2005). Alpha-synuclein cooperates with CSPalpha in preventing

Chiti, F.&C. M. Dobson (2006). Protein misfolding, functional amyloid, and human disease.

Choi, W., et al. (2004). Mutation E46K increases phospholipid binding and assembly into

Christine, C. W., et al. (2009). Safety and tolerability of putaminal AADC gene therapy for

Chung, K. K., et al. (2001). Parkin ubiquitinates the alpha-synuclein-interacting protein,

Danzer, K. M., et al. (2011). Heat-shock protein 70 modulates toxic extracellular alpha-

Danzer, K. M., et al. (2011). Heat-shock protein 70 modulates toxic extracellular alpha-

De Mena, L., et al. (2009). Mutational screening of the mortalin gene (HSPA9) in Parkinson's

Dedmon, M. M., et al. (2005). Heat shock protein 70 inhibits alpha-synuclein fibril formation via preferential binding to prefibrillar species. *J Biol Chem*, 280, (15), pp. (14733-40). Demand, J., et al. (2001). Cooperation of a ubiquitin domain protein and an E3 ubiquitin ligase during chaperone/proteasome coupling. *Curr Biol*, 11, (20), pp. (1569-77).

synphilin-1: implications for Lewy-body formation in Parkinson disease. *Nat Med*,

synuclein oligomers and rescues trans-synaptic toxicity. *FASEB J*, 25, (1), pp. (326-

synuclein oligomers and rescues trans-synaptic toxicity. *FASEB J*, 25, (1), pp. (326-

filaments of human alpha-synuclein. *FEBS Lett*, 576, (3), pp. (363-8).

Parkinson's disease. *Biochim Biophys Acta*, 1792, (7), pp. (664-75).

mediated autophagy. *Science*, 305, (5688), pp. (1292-5).

neurodegeneration. *Cell*, 123, (3), pp. (383-96).

Parkinson disease. *Neurology*, 73, (20), pp. (1662-9).

disease. *J Neural Transm*, 116, (10), pp. (1289-93).

*Annu Rev Biochem*, 75, pp. (333-66).

7, (10), pp. (1144-50).

36).

36).

neuron loss in Parkinson's models. *Science*, 313, (5785), pp. (324-8).

homeostasis. *Hum Mol Genet*, 19, (22), pp. (4437-52).

by heat-shock proteins. *Nat Cell Biol*, 3, (1), pp. (93-6).

pp. (571-6).

Parkinson's disease: functional impact of disease-related variants on mitochondrial

property of both alpha-synuclein mutations linked to early-onset Parkinson's disease: implications for pathogenesis and therapy. *Proc Natl Acad Sci U S A*, 97, (2),

#### **7. References**


Adachi, H., et al. (2007). CHIP overexpression reduces mutant androgen receptor protein

Ahmad, A. (2010). DnaK/DnaJ/GrpE of Hsp70 system have differing effects on alpha-

Al-Ramahi, I., et al. (2006). CHIP protects from the neurotoxicity of expanded and wild-type

Alberti, S., et al. (2004). The cochaperone HspBP1 inhibits the CHIP ubiquitin ligase and

Alberti, S., et al. (2002). Ubiquitylation of BAG-1 suggests a novel regulatory mechanism

Alvarez-Erviti, L., et al. (2010). Chaperone-mediated autophagy markers in Parkinson

Arawaka, S., et al. (2010). Heat shock proteins as suppressors of accumulation of toxic

Auluck, P. K., et al. (2002). Chaperone suppression of alpha-synuclein toxicity in a Drosophila model for Parkinson's disease. *Science*, 295, (5556), pp. (865-8). Auluck, P. K., et al. (2005). Mechanisms of Suppression of {alpha}-Synuclein Neurotoxicity

Ballinger, C. A., et al. (1999). Identification of CHIP, a novel tetratricopeptide repeat-

Bandopadhyay, R.&J. de Belleroche (2010). Pathogenesis of Parkinson's disease: emerging

Bandyopadhyay, U.&A. M. Cuervo (2007). Chaperone-mediated autophagy in aging and

Bellucci, A., et al. (2011). Induction of the unfolded protein response by alpha-synuclein in experimental models of Parkinson's disease. *J Neurochem*, 116, (4), pp. (588-605). Bence, N. F., et al. (2001). Impairment of the ubiquitin-proteasome system by protein

Bercovich, B., et al. (1997). Ubiquitin-dependent degradation of certain protein substrates in vitro requires the molecular chaperone Hsc70. *J Biol Chem*, 272, (14), pp. (9002-10). Braun, J. E., et al. (1996). The cysteine string secretory vesicle protein activates Hsc70

Broadley, S. A.&F. U. Hartl (2009). The role of molecular chaperones in human misfolding

Bukau, B.&A. L. Horwich (1998). The Hsp70 and Hsp60 chaperone machines. *Cell*, 92, (3),

by Geldanamycin in Drosophila. *J Biol Chem*, 280, (4), pp. (2873-8).

role of molecular chaperones. *Trends Mol Med*, 16, (1), pp. (27-36).

chaperone functions. *Mol Cell Biol*, 19, (6), pp. (4535-45).

aggregation. *Science*, 292, (5521), pp. (1552-5).

ATPase. *J Biol Chem*, 271, (42), pp. (25989-93).

diseases. *FEBS Lett*, 583, (16), pp. (2647-53).

mouse model. *J Neurosci*, 27, (19), pp. (5115-26).

regulator. *Mol Biol Cell*, 15, (9), pp. (4003-10).

disease brains. *Arch Neurol*, 67, (12), pp. (1464-72).

*Curr Pharm Biotechnol*, 11, (2), pp. (158-66).

and ameliorates phenotypes of the spinal and bulbar muscular atrophy transgenic

synuclein fibrillation involved in Parkinson's disease. *Int J Biol Macromol*, 46, (2), pp.

ataxin-1 and promotes their ubiquitination and degradation. *J Biol Chem*, 281, (36),

stimulates the maturation of the cystic fibrosis transmembrane conductance

during the sorting of chaperone substrates to the proteasome. *J Biol Chem*, 277, (48),

prefibrillar intermediates and misfolded proteins in neurodegenerative diseases.

containing protein that interacts with heat shock proteins and negatively regulates

neurodegeneration: lessons from alpha-synuclein. *Exp Gerontol*, 42, (1-2), pp. (120-

**7. References** 

(275-9).

pp. (26714-24).

pp. (45920-7).

8).

pp. (351-66).


The Hsp70 Chaperone System in Parkinson's Disease 239

Grunblatt, E., et al. (2001). Gene expression analysis in N-methyl-4-phenyl-1,2,3,6-

Grunblatt, E., et al. (2010). Pilot study: peripheral biomarkers for diagnosing sporadic

Gupta, A., et al. (2008). What causes cell death in Parkinson's disease? *Ann Neurol*, 64 Suppl

Hartl, F. U. (1996). Molecular chaperones in cellular protein folding. *Nature*, 381, (6583), pp.

Hartl, F. U.&M. Hayer-Hartl (2002). Molecular chaperones in the cytosol: from nascent chain

Hartl, F. U.&M. Hayer-Hartl (2009). Converging concepts of protein folding in vitro and in

Hasegawa, M., et al. (2002). Phosphorylated alpha-synuclein is ubiquitinated in alpha-

Hatakeyama, S., et al. (2004). U-box protein carboxyl terminus of Hsc70-interacting protein

Hershko, A.&A. Ciechanover (1998). The ubiquitin system. *Annu Rev Biochem*, 67, pp. (425-

Hinault, M. P., et al. (2010). Stable alpha-synuclein oligomers strongly inhibit chaperone

Hohfeld, J., et al. (2001). From the cradle to the grave: molecular chaperones that may choose

Hohfeld, J., et al. (1995). Hip, a novel cochaperone involved in the eukaryotic Hsc70/Hsp40

Hong, Z., et al. (2010). DJ-1 and alpha-synuclein in human cerebrospinal fluid as biomarkers

Hoozemans, J. J., et al. (2007). Activation of the unfolded protein response in Parkinson's

Hou, Y.&J. Zou (2009). Delivery of HSF1(+) protein using HIV-1 TAT protein transduction

Huang, C., et al. (2006). Heat shock protein 70 inhibits alpha-synuclein fibril formation via interactions with diverse intermediates. *J Mol Biol*, 364, (3), pp. (323-36). Imai, Y., et al. (2002). CHIP is associated with Parkin, a gene responsible for familial

Imai, Y., et al. (2001). An unfolded putative transmembrane polypeptide, which can lead to endoplasmic reticulum stress, is a substrate of Parkin. *Cell*, 105, (7), pp. (891-902). Jellinger, K. A. (2010). Basic mechanisms of neurodegeneration: a critical update. *J Cell Mol* 

Parkinson's disease, and enhances its ubiquitin ligase activity. *Mol Cell*, 10, (1), pp.

between folding and degradation. *EMBO Rep*, 2, (10), pp. (885-90).

(CHIP) mediates poly-ubiquitylation preferentially on four-repeat Tau and is involved in neurodegeneration of tauopathy. *J Neurochem*, 91, (2), pp. (299-307). Hauser, M. A., et al. (2005). Expression profiling of substantia nigra in Parkinson disease,

progressive supranuclear palsy, and frontotemporal dementia with parkinsonism.

activity of the Hsp70 system by weak interactions with J-domain co-chaperones. *J* 

effect of R-apomorphine. *J Neurochem*, 78, (1), pp. (1-12).

to folded protein. *Science*, 295, (5561), pp. (1852-8).

vivo. *Nat Struct Mol Biol*, 16, (6), pp. (574-81).

*Arch Neurol*, 62, (6), pp. (917-21).

*Biol Chem*, 285, (49), pp. (38173-82).

reaction cycle. *Cell*, 83, (4), pp. (589-98).

domain. *Mol Biol Rep*, 36, (8), pp. (2271-7).

of Parkinson's disease. *Brain*, 133, (Pt 3), pp. (713-26).

disease. *Biochem Biophys Res Commun*, 354, (3), pp. (707-11).

2, pp. (S3-15).

(571-9).

79).

(55-67).

*Med*, 14, (3), pp. (457-87).

Parkinson's disease. *J Neural Transm*, 117, (12), pp. (1387-93).

synucleinopathy lesions. *J Biol Chem*, 277, (50), pp. (49071-6).

tetrahydropyridine mice model of Parkinson's disease using cDNA microarray:


Devi, L., et al. (2008). Mitochondrial import and accumulation of alpha-synuclein impair

Dice, J. F. (1990). Peptide sequences that target cytosolic proteins for lysosomal proteolysis.

Dickey, C. A., et al. (2007). Brain CHIP: removing the culprits in neurodegenerative disease.

Dong, Z., et al. (2005). Hsp70 gene transfer by adeno-associated virus inhibits MPTP-

Dul, J. L., et al. (2001). Hsp70 and antifibrillogenic peptides promote degradation and inhibit

El-Agnaf, O. M., et al. (1998). Aggregates from mutant and wild-type alpha-synuclein

Etminan, M., et al. (2008). Non-steroidal anti-inflammatory drug use and the risk of Parkinson disease: a retrospective cohort study. *J Clin Neurosci*, 15, (5), pp. (576-7). Fasano, M., et al. (2008). Peripheral biomarkers of Parkinson's disease as early reporters of

Fawell, S., et al. (1994). Tat-mediated delivery of heterologous proteins into cells. *Proc Natl* 

Fornai, F., et al. (2005). Parkinson-like syndrome induced by continuous MPTP infusion:

Fujikake, N., et al. (2008). Heat shock transcription factor 1-activating compounds suppress

Gagne, J. J.&M. C. Power (2010). Anti-inflammatory drugs and risk of Parkinson disease: a

Garcia-Mata, R., et al. (1999). Characterization and dynamics of aggresome formation by a

Garcia-Reitbock, P., et al. (2010). SNARE protein redistribution and synaptic failure in a transgenic mouse model of Parkinson's disease. *Brain*, 133, (Pt 7), pp. (2032-44). Giasson, B. I.&V. M. Lee (2003). Are ubiquitination pathways central to Parkinson's disease?

Giasson, B. I., et al. (2001). A hydrophobic stretch of 12 amino acid residues in the middle of

Gibbs, S. J., et al. (2009). Hsp40 couples with the CSPalpha chaperone complex upon

Gosavi, N., et al. (2002). Golgi fragmentation occurs in the cells with prefibrillar alpha-

induction of the heat shock response. *PLoS One*, 4, (2), pp. (e4595).

alpha-synuclein is essential for filament assembly. *J Biol Chem*, 276, (4), pp. (2380-6).

synuclein aggregates and precedes the formation of fibrillar inclusion. *J Biol Chem*,

convergent roles of the ubiquitin-proteasome system and alpha-synuclein. *Proc Natl* 

polyglutamine-induced neurodegeneration through induction of multiple

central neurodegeneration. *Biomark Med*, 2, (5), pp. (465-78).

molecular chaperones. *J Biol Chem*, 283, (38), pp. (26188-97).

cytosolic GFP-chimera. *J Cell Biol*, 146, (6), pp. (1239-54).

meta-analysis. *Neurology*, 74, (12), pp. (995-1002).

Dice, J. F. (2007). Chaperone-mediated autophagy. *Autophagy*, 3, (4), pp. (295-9).

*Biol Chem*, 283, (14), pp. (9089-100).

*Trends Mol Med*, 13, (1), pp. (32-8).

*Acad Sci U S A*, 91, (2), pp. (664-8).

*Acad Sci U S A*, 102, (9), pp. (3413-8).

*Cell*, 114, (1), pp. (1-8).

277, (50), pp. (48984-92).

*Ther*, 11, (1), pp. (80-8).

16).

pp. (71-5).

*Trends Biochem Sci*, 15, (8), pp. (305-9).

complex I in human dopaminergic neuronal cultures and Parkinson disease brain. *J* 

induced nigrostriatal degeneration in the mouse model of Parkinson disease. *Mol* 

intracellular aggregation of amyloidogenic light chains. *J Cell Biol*, 152, (4), pp. (705-

proteins and NAC peptide induce apoptotic cell death in human neuroblastoma cells by formation of beta-sheet and amyloid-like filaments. *FEBS Lett*, 440, (1-2),


The Hsp70 Chaperone System in Parkinson's Disease 241

Lee, B. S., et al. (1995). Pharmacological modulation of heat shock factor 1 by

Leroy, E., et al. (1998). The ubiquitin pathway in Parkinson's disease. *Nature*, 395, (6701), pp.

Liani, E., et al. (2004). Ubiquitylation of synphilin-1 and alpha-synuclein by SIAH and its

Lindersson, E., et al. (2004). Proteasomal inhibition by alpha-synuclein filaments and

Liu, Y., et al. (2002). The UCH-L1 gene encodes two opposing enzymatic activities that affect

Lo Bianco, C., et al. (2008). Hsp104 antagonizes alpha-synuclein aggregation and reduces

Lotz, G. P., et al. (2010). Hsp70 and Hsp40 functionally interact with soluble mutant

Lu, T. Z., et al. (2010). Multifaceted role of heat shock protein 70 in neurons. *Mol Neurobiol*,

Luders, J., et al. (2000). The ubiquitin-related BAG-1 provides a link between the molecular chaperones Hsc70/Hsp70 and the proteasome. *J Biol Chem*, 275, (7), pp. (4613-7). Luk, K. C., et al. (2008). Interactions between Hsp70 and the hydrophobic core of alphasynuclein inhibit fibril assembly. *Biochemistry*, 47, (47), pp. (12614-25). Maeda, H., et al. (2007). Biological heterogeneity of the peptide-binding motif of the 70-kDa

Mak, S. K., et al. (2010). Lysosomal degradation of alpha-synuclein in vivo. *J Biol Chem*, 285,

Manfredsson, F. P., et al. (2006). RNA knockdown as a potential therapeutic strategy in

Manning-Bog, A. B., et al. (2006). Lack of nigrostriatal pathology in a rat model of

Martinez-Vicente, M., et al. (2008). Dopamine-modified alpha-synuclein blocks chaperone-

Martinez-Vicente, M., et al. Cargo recognition failure is responsible for inefficient autophagy

Mayer, M. P.&B. Bukau (2005). Hsp70 chaperones: cellular functions and molecular

McDonough, H.&C. Patterson (2003). CHIP: a link between the chaperone and proteasome

McLean, P. J., et al. (2002). TorsinA and heat shock proteins act as molecular chaperones: suppression of alpha-synuclein aggregation. *J Neurochem*, 83, (4), pp. (846-54). McLean, P. J., et al. (2004). Geldanamycin induces Hsp70 and prevents alpha-synuclein

aggregation and toxicity in vitro. *Biochem Biophys Res Commun*, 321, (3), pp. (665-9).

Parkinson's disease. *Gene Ther*, 13, (6), pp. (517-24).

mechanism. *Cell Mol Life Sci*, 62, (6), pp. (670-84).

systems. *Cell Stress Chaperones*, 8, (4), pp. (303-8).

proteasome inhibition. *Ann Neurol*, 60, (2), pp. (256-60).

mediated autophagy. *J Clin Invest*, 118, (2), pp. (777-88).

in Huntington's disease. *Nat Neurosci*, 13, (5), pp. (567-76).

damage. *Proc Natl Acad Sci U S A*, 92, (16), pp. (7207-11).

*Proc Natl Acad Sci U S A*, 101, (15), pp. (5500-5).

oligomers. *J Biol Chem*, 279, (13), pp. (12924-34).

(451-2).

pp. (209-18).

(9), pp. (3087-97).

(49), pp. (38183-93).

42, (2), pp. (114-23).

(26956-62).

(18), pp. (13621-9).

antiinflammatory drugs results in protection against stress-induced cellular

presence in cellular inclusions and Lewy bodies imply a role in Parkinson's disease.

alpha-synuclein degradation and Parkinson's disease susceptibility. *Cell*, 111, (2),

dopaminergic degeneration in a rat model of Parkinson disease. *J Clin Invest*, 118,

huntingtin oligomers in a classic ATP-dependent reaction cycle. *J Biol Chem*, 285,

heat shock protein by surface plasmon resonance analysis. *J Biol Chem*, 282, (37), pp.


Jin, J., et al. (2006). Proteomic identification of a stress protein, mortalin/mthsp70/GRP75: relevance to Parkinson disease. *Mol Cell Proteomics*, 5, (7), pp. (1193-204). Jin, J., et al. (2007). Identification of novel proteins associated with both alpha-synuclein and

Johnson, J. N., et al. (2010). CSPalpha: the neuroprotective J protein. *Biochem Cell Biol*, 88, (2),

Jung, A. E., et al. (2008). HSP70 and constitutively active HSF1 mediate protection against

Jurivich, D. A., et al. (1995). Salicylate triggers heat shock factor differently than heat. *J Biol* 

Kalia, L. V., et al. (2011). Ubiquitinylation of alpha-Synuclein by Carboxyl Terminus Hsp70-

Kalia, S. K., et al. (2004). BAG5 inhibits parkin and enhances dopaminergic neuron

Kampinga, H. H. & E. A. Craig (2010). The HSP70 chaperone machinery: J proteins as drivers of functional specificity. *Nat Rev Mol Cell Biol*, 11, (8), pp. (579-92). Karpinar, D. P., et al. (2009). Pre-fibrillar alpha-synuclein variants with impaired beta-

Kim, S. A., et al. (2008). TAT-Hsp40 inhibits oxidative stress-mediated cytotoxicity via the

Kitada, T., et al. (1998). Mutations in the parkin gene cause autosomal recessive juvenile

Klucken, J., et al. (2006). Detection of novel intracellular alpha-synuclein oligomeric species

Klucken, J., et al. (2004a). A single amino acid substitution differentiates Hsp70-dependent

Klucken, J., et al. (2004b). Hsp70 Reduces alpha-Synuclein Aggregation and Toxicity. *J Biol* 

Ko, H. S., et al. (2009). CHIP regulates leucine-rich repeat kinase-2 ubiquitination, degradation, and toxicity. *Proc Natl Acad Sci U S A*, 106, (8), pp. (2897-902). Koga, H.&A. M. Cuervo (2010). Chaperone-mediated autophagy dysfunction in the

Kruger, R., et al. (1998). Ala30Pro mutation in the gene encoding alpha-synuclein in

Lai, Y., et al. (2005). Selectively increasing inducible heat shock protein 70 via TAT-protein

Lashuel, H. A., et al. (2002a). Neurodegenerative disease: amyloid pores from pathogenic

Lashuel, H. A., et al. (2002b). Alpha-synuclein, especially the Parkinson's disease-associated

transduction protects neurons from nitrosative stress and excitotoxicity. *J* 

mutants, forms pore-like annular and tubular protofibrils. *J Mol Biol*, 322, (5), pp.

effects on alpha-synuclein degradation and toxicity. *Biochem Biophys Res Commun*,

inhibition of Hsp70 ubiquitination. *FEBS Lett*, 582, (5), pp. (734-40).

by fluorescence lifetime imaging. *FASEB J*, 20, (12), pp. (2050-7).

pathogenesis of neurodegeneration. *Neurobiol Dis*, pp.

Parkinson's disease. *Nat Genet*, 18, (2), pp. (106-8).

Interacting Protein (CHIP) Is Regulated by Bcl-2-Associated Athanogene 5 (BAG5).

structure increase neurotoxicity in Parkinson's disease models. *EMBO J*, 28, (20),

CDCrel-1-mediated toxicity. *Mol Ther*, 16, (6), pp. (1048-55).

DJ-1. *Mol Cell Proteomics*, 6, (5), pp. (845-59).

degeneration. *Neuron*, 44, (6), pp. (931-45).

parkinsonism. *Nature*, 392, (6676), pp. (605-8).

*Chem*, 270, (41), pp. (24489-95).

*PLoS One*, 6, (2), pp. (e14695).

pp. (157-65).

pp. (3256-68).

325, (1), pp. (367-73).

*Chem*, 279, (24), pp. (25497-502).

*Neurochem*, 94, (2), pp. (360-6).

(1089-102).

mutations. *Nature*, 418, (6895), pp. (291).


The Hsp70 Chaperone System in Parkinson's Disease 243

Pacey, S., et al. (2010). A Phase II trial of 17-allylamino, 17-demethoxygeldanamycin (17- AAG, tanespimycin) in patients with metastatic melanoma. *Invest New Drugs*, pp. Paschen, W.&T. Mengesdorf (2005). Endoplasmic reticulum stress response and

Pennington, K., et al. (2010). Differential effects of wild-type and A53T mutant isoform of

Petrucelli, L., et al. (2002). Parkin protects against the toxicity associated with mutant alpha-

Phukan, J. (2010). Arimoclomol, a coinducer of heat shock proteins for the potential treatment of amyotrophic lateral sclerosis. *IDrugs*, 13, (7), pp. (482-96). Polymeropoulos, M. H., et al. (1997). Mutation in the alpha-synuclein gene identified in

Prapapanich, V., et al. (1996). Molecular cloning of human p48, a transient component of

Putcha, P., et al. (2010). Brain-permeable small-molecule inhibitors of Hsp90 prevent alpha-

Richardson, P. G., et al. (2010). Tanespimycin with bortezomib: activity in

Rideout, H. J., et al. (2005). Dopaminergic neurons in rat ventral midbrain cultures undergo

Riedel, M., et al. (2010). 17-AAG induces cytoplasmic alpha-synuclein aggregate clearance

Rochet, J. C., et al. (2000). Inhibition of fibrillization and accumulation of prefibrillar

Roodveldt, C., et al. (2009). Chaperone proteostasis in Parkinson's disease: stabilization of the Hsp70/alpha-synuclein complex by Hip. *EMBO J*, 28, (23), pp. (3758-70). Roodveldt, C., et al. (2008). Immunological features of alpha-synuclein in Parkinson's

Rosales-Hernandez, A., et al. (2009). RDJ2 (DNAJA2) chaperones neural G protein signaling

Rott, R., et al. (2008). Monoubiquitylation of alpha-synuclein by seven in absentia homolog

Rubinsztein, D. C. (2006). The roles of intracellular protein-degradation pathways in

Rudiger, S., et al. (1997). Substrate specificity of the DnaK chaperone determined by screening cellulose-bound peptide libraries. *EMBO J*, 16, (7), pp. (1501-7). Rutkowski, D. T.&R. J. Kaufman (2004). A trip to the ER: coping with stress. *Trends Cell Biol*,

(SIAH) promotes its aggregation in dopaminergic cells. *J Biol Chem*, 283, (6), pp.

proteasomal inhibition. *J Neurochem*, 93, (5), pp. (1304-13).

by induction of autophagy. *PLoS One*, 5, (1), pp. (e8753).

disease. *J Cell Mol Med*, 12, (5B), pp. (1820-9).

pathways. *Cell Stress Chaperones*, 14, (1), pp. (71-82).

neurodegeneration. *Nature*, 443, (7113), pp. (780-6).

families with Parkinson's disease. *Science*, 276, (5321), pp. (2045-7).

alpha-synuclein on the mitochondrial proteome of differentiated SH-SY5Y cells. *J* 

synuclein: proteasome dysfunction selectively affects catecholaminergic neurons.

progesterone receptor complexes and an Hsp70-binding protein. *Mol Endocrinol*, 10,

synuclein oligomer formation and rescue alpha-synuclein-induced toxicity. *J* 

relapsed/refractory patients with multiple myeloma. *Br J Haematol*, 150, (4), pp.

selective apoptosis and form inclusions, but do not up-regulate iHSP70, following

oligomers in mixtures of human and mouse alpha-synuclein. *Biochemistry*, 39, (35),

neurodegeneration. *Cell Calcium*, 38, (3-4), pp. (409-15).

*Proteome Res*, 9, (5), pp. (2390-401).

*Pharmacol Exp Ther*, 332, (3), pp. (849-57).

*Neuron*, 36, (6), pp. (1007-19).

(4), pp. (420-31).

(428-37).

pp. (10619-26).

(3316-28).

14, (1), pp. (20-8).


McNaught, K. S., et al. (2003). Altered proteasomal function in sporadic Parkinson's disease.

McNaught, K. S.&P. Jenner (2001). Proteasomal function is impaired in substantia nigra in

McNaught, K. S., et al. (2002a). Impairment of the ubiquitin-proteasome system causes

McNaught, K. S.&C. W. Olanow (2006). Proteasome inhibitor-induced model of Parkinson's

McNaught, K. S., et al. (2001). Failure of the ubiquitin-proteasome system in Parkinson's

McNaught, K. S., et al. (2004). Systemic exposure to proteasome inhibitors causes a progressive model of Parkinson's disease. *Ann Neurol*, 56, (1), pp. (149-62). McNaught, K. S., et al. (2002b). Aggresome-related biogenesis of Lewy bodies. *Eur J* 

Meacham, G. C., et al. (2001). The Hsc70 co-chaperone CHIP targets immature CFTR for

Miller, L. C., et al. (2003). Cysteine string protein (CSP) inhibition of N-type calcium channels is blocked by mutant huntingtin. *J Biol Chem*, 278, (52), pp. (53072-81). Minami, Y., et al. (1996). Regulation of the heat-shock protein 70 reaction cycle by the mammalian DnaJ homolog, Hsp40. *J Biol Chem*, 271, (32), pp. (19617-24). Morgan, J. C., et al. (2010). Biomarkers in Parkinson's disease. *Curr Neurol Neurosci Rep*, 10,

Morimoto, R. I. (2008). Proteotoxic stress and inducible chaperone networks in neurodegenerative disease and aging. *Genes Dev*, 22, (11), pp. (1427-38). Mouradian, M. M. (2002). Recent advances in the genetics and pathogenesis of Parkinson

Muchowski, P. J., et al. (2000). Hsp70 and hsp40 chaperones can inhibit self-assembly of

Murata, S., et al. (2001). CHIP is a chaperone-dependent E3 ligase that ubiquitylates

Nagel, F., et al. (2008). Tat-Hsp70 protects dopaminergic neurons in midbrain cultures and

Neef, D. W., et al. (2010). Modulation of heat shock transcription factor 1 as a therapeutic

Nixon, R. A. (2006). Autophagy in neurodegenerative disease: friend, foe or turncoat? *Trends* 

Nyhlen, J., et al. (2010). Problems associated with fluid biomarkers for Parkinson's disease.

Opazo, F., et al. (2008). Accumulation and clearance of alpha-synuclein aggregates demonstrated by time-lapse imaging. *J Neurochem*, 106, (2), pp. (529-40). Outeiro, T. F., et al. (2008). Formation of toxic oligomeric alpha-synuclein species in living

polyglutamine proteins into amyloid-like fibrils. *Proc Natl Acad Sci U S A*, 97, (14),

in the substantia nigra in models of Parkinson's disease. *J Neurochem*, 105, (3), pp.

target for small molecule intervention in neurodegenerative disease. *PLoS Biol*, 8,

dopaminergic cell death and inclusion body formation in ventral mesencephalic

Parkinson's disease. *Neurosci Lett*, 297, (3), pp. (191-4).

proteasomal degradation. *Nat Cell Biol*, 3, (1), pp. (100-5).

*Exp Neurol*, 179, (1), pp. (38-46).

cultures. *J Neurochem*, 81, (2), pp. (301-6).

disease. *Ann Neurol*, 60, (2), pp. (243-7).

disease. *Neurology*, 58, (2), pp. (179-85).

unfolded protein. *EMBO Rep*, 2, (12), pp. (1133-8).

*Neurosci*, 16, (11), pp. (2136-48).

(6), pp. (423-30).

pp. (7841-6).

(853-64).

(1), pp. (e1000291).

*Neurosci*, 29, (9), pp. (528-35).

*Biomark Med*, 4, (5), pp. (671-81).

cells. *PLoS One*, 3, (4), pp. (e1867).

disease. *Nat Rev Neurosci*, 2, (8), pp. (589-94).


The Hsp70 Chaperone System in Parkinson's Disease 245

Tobaben, S., et al. (2001). A trimeric protein complex functions as a synaptic chaperone

Urushitani, M., et al. (2004). CHIP promotes proteasomal degradation of familial ALS-linked mutant SOD1 by ubiquitinating Hsp/Hsc70. *J Neurochem*, 90, (1), pp. (231-44). Valente, E. M., et al. (2004). Hereditary early-onset Parkinson's disease caused by mutations

Vashist, S., et al. (2010). Applying Hsp104 to protein-misfolding disorders. *Biochem Cell Biol*,

Vila, M.&S. Przedborski (2003). Targeting programmed cell death in neurodegenerative

Vogiatzi, T., et al. (2008). Wild type alpha-synuclein is degraded by chaperone-mediated

Voisine, C., et al. (2010). Chaperone networks: tipping the balance in protein folding

Volles, M. J.&P. T. Lansbury, Jr. (2002). Vesicle permeabilization by protofibrillar alpha-

Volles, M. J.&P. T. Lansbury, Jr. (2003). Zeroing in on the pathogenic form of alpha-

Wacker, J. L., et al. (2004). Hsp70 and Hsp40 attenuate formation of spherical and annular

Waragai, M., et al. (2007). Plasma levels of DJ-1 as a possible marker for progression of

Warrick, J. M., et al. (1999). Suppression of polyglutamine-mediated neurodegeneration in Drosophila by the molecular chaperone HSP70. *Nat Genet*, 23, (4), pp. (425-8). Waza, M., et al. (2005). 17-AAG, an Hsp90 inhibitor, ameliorates polyglutamine-mediated

Waza, M., et al. (2006). Modulation of Hsp90 function in neurodegenerative disorders: a

Westerheide, S. D., et al. (2004). Celastrols as inducers of the heat shock response and

Witt, S. N. (2009). Hsp70 molecular chaperones and Parkinson's disease. *Biopolymers*, 93, (3),

Wong, E.&A. M. Cuervo (2010). Autophagy gone awry in neurodegenerative diseases. *Nat* 

Wood, S. J., et al. (1999). alpha-synuclein fibrillogenesis is nucleation-dependent.

Xie, W., et al. (2010). New insights into the role of mitochondrial dysfunction and protein aggregation in Parkinson's disease. *Biochim Biophys Acta*, 1802, (11), pp. (935-41). Xilouri, M.&L. Stefanis (2011). Autophagic pathways in Parkinson disease and related

Implications for the pathogenesis of Parkinson's disease. *J Biol Chem*, 274, (28), pp.

molecular-targeted therapy against disease-causing protein. *J Mol Med*, 84, (8), pp.

sporadic Parkinson's disease. *Neurosci Lett*, 425, (1), pp. (18-22).

motor neuron degeneration. *Nat Med*, 11, (10), pp. (1088-95).

cytoprotection. *J Biol Chem*, 279, (53), pp. (56053-60).

disorders. *Expert Rev Mol Med*, 13, pp. (e8).

autophagy and macroautophagy in neuronal cells. *J Biol Chem*, 283, (35), pp. (23542-

synuclein is sensitive to Parkinson's disease-linked mutations and occurs by a pore-

synuclein and its mechanism of neurotoxicity in Parkinson's disease. *Biochemistry*,

polyglutamine oligomers by partitioning monomer. *Nat Struct Mol Biol*, 11, (12), pp.

machine. *Neuron*, 31, (6), pp. (987-99).

88, (1), pp. (1-13).

42, (26), pp. (7871-8).

(1215-22).

(635-46).

pp. (218-28).

(19509-12).

*Neurosci*, 13, (7), pp. (805-11).

56).

in PINK1. *Science*, 304, (5674), pp. (1158-60).

diseases. *Nat Rev Neurosci*, 4, (5), pp. (365-75).

diseases. *Neurobiol Dis*, 40, (1), pp. (12-20).

like mechanism. *Biochemistry*, 41, (14), pp. (4595-602).


Sakisaka, T., et al. (2002). Rab-alphaGDI activity is regulated by a Hsp90 chaperone

Sato, S., et al. (2006). 14-3-3eta is a novel regulator of parkin ubiquitin ligase. *EMBO J*, 25, (1),

Scherzer, C. R., et al. (2007). Molecular markers of early Parkinson's disease based on gene

Scheufler, C., et al. (2000). Structure of TPR domain-peptide complexes: critical elements in

Schlecht, R., et al. (2011). Mechanics of Hsp70 chaperones enables differential interaction

Schwarze, S. R., et al. (1999). In vivo protein transduction: delivery of a biologically active

Shadrina, M. I., et al. (2010). Expression analysis of suppression of tumorigenicity 13 gene in

Sharma, M., et al. (2011). CSPalpha promotes SNARE-complex assembly by chaperoning

Shen, H. Y., et al. (2005). Geldanamycin induces heat shock protein 70 and protects against

Shi, M., et al. (2010). Significance and confounders of peripheral DJ-1 and alpha-synuclein in

Shimshek, D. R., et al. (2010). The HSP70 molecular chaperone is not beneficial in a mouse

Shimura, H., et al. (2001). Ubiquitination of a new form of alpha-synuclein by parkin from human brain: implications for Parkinson's disease. *Science*, 293, (5528), pp. (263-9). Shin, Y., et al. (2005). The co-chaperone carboxyl terminus of Hsp70-interacting protein

Singleton, A. B., et al. (2003). alpha-Synuclein locus triplication causes Parkinson's disease.

Solit, D. B., et al. (2008). Phase II trial of 17-allylamino-17-demethoxygeldanamycin in patients with metastatic melanoma. *Clin Cancer Res*, 14, (24), pp. (8302-7). Spillantini, M. G., et al. (1998). alpha-Synuclein in filamentous inclusions of Lewy bodies

Sun, L.&Z. J. Chen (2004). The novel functions of ubiquitination in signaling. *Curr Opin Cell* 

Takayama, S., et al. (1999). An evolutionarily conserved family of Hsp70/Hsc70 molecular

Taxis, C., et al. (2003). Use of modular substrates demonstrates mechanistic diversity and

Tetzlaff, J. E., et al. (2008). CHIP targets toxic alpha-Synuclein oligomers for degradation. *J* 

MPTP-induced dopaminergic neurotoxicity in mice. *J Biol Chem*, 280, (48), pp.

(CHIP) mediates alpha-synuclein degradation decisions between proteasomal and

from Parkinson's disease and dementia with lewy bodies. *Proc Natl Acad Sci U S A*,

reveals differences in chaperone requirement of ERAD. *J Biol Chem*, 278, (38), pp.

patients with Parkinson's disease. *Neurosci Lett*, 473, (3), pp. (257-9).

SNAP-25 during synaptic activity. *Nat Cell Biol*, 13, (1), pp. (30-9).

model of alpha-synucleinopathy. *PLoS One*, 5, (4), pp. (e10014).

lysosomal pathways. *J Biol Chem*, 280, (25), pp. (23727-34). Shults, C. W. (2006). Lewy bodies. *Proc Natl Acad Sci U S A*, 103, (6), pp. (1661-8).

chaperone regulators. *J Biol Chem*, 274, (2), pp. (781-6).

*Science*, 302, (5646), pp. (841).

95, (11), pp. (6469-73).

(35903-13).

*Biol*, 16, (2), pp. (119-26).

*Biol Chem*, 283, (26), pp. (17962-8).

the assembly of the Hsp70-Hsp90 multichaperone machine. *Cell*, 101, (2), pp. (199-

expression in blood. *Proc Natl Acad Sci U S A*, 104, (3), pp. (955-60).

with client proteins. *Nat Struct Mol Biol*, 18, (3), pp. (345-51).

protein into the mouse. *Science*, 285, (5433), pp. (1569-72).

Parkinson's disease. *Neurosci Lett*, 480, (1), pp. (78-82).

complex. *EMBO J*, 21, (22), pp. (6125-35).

pp. (211-21).

210).

(39962-9).


**11** 

*USA* 

**The Noradrenergic System is a Major** 

**Component in Parkinson's Disease** 

Patricia Szot1, Allyn Franklin1 and Murray A. Raskind1,2

*1Veterans Administration Northwest Network for Mental Illness Research, Education and Clinical Center and Puget Sound Health Care System, Seattle,* 

*2Department of Psychiatry and Behavioral Science, University of Washington, Seattle* 

Parkinson's disease (PD) is a neurological disorder that affects approximately 2% of the elderly population, and as our population continues to age, the incidence will only increase (Singh et al., 2007). PD is commonly characterized by various motor deficits including tremor, rigidity and bradykinesia (Singh et al., 2007). The cause of these motor symptoms is the loss of dopaminergic neurons in the substantia nigra pars compacta (SN) and reduced dopamine (DA) levels in the striatum (Damier et al., 1999; Gibb, 1991; Gibb and Lee, 1991). However, the appearance of PD symptoms does not occur until 70-80% of the dopaminergic neurons are lost. In the progression of this disorder, the loss of dopaminergic neurons is not observed until Stage 3 (out of 6 Stages) of the disorder (Braak et al., 2003a, 2003b, 2006).

Of course a great deal of research has focused on the dopaminergic system in PD because loss of neurons in the SN is responsible for PD symptoms; however, PD is represented by multiple systems failing. During the earlier stages of the disorder, non-motor preclinical symptoms are observed. These preclinical symptoms include hyposmia (Berendse et al., 2001, Ponsen et al., 2004), REM-sleep disorder (Boeve et al., 2003, Scheneck et al., 2003), depression (Leentjens et al., 2003; Mayeux et al., 1992; Slaughter et al., 2001) and autonomic dysfunction such as orthostatic hypotension (Mathias, 1998; Ziemssen & Reichmann, 2007). These preclinical symptoms are attributed to neuropathological changes in neurotransmitter systems other than the SN dopaminergic nervous system. One neurotransmitter system that may be responsible for these early non-motor symptoms is the noradrenergic nervous system (Goldstein et al., 2011; Itoi & Sugimoto, 2010; Lopez-Munoz & Alamo, 2009; Mathias, 1998; Osaka & Matsumura, 1994; Ziegler et al., 1977). The presence of these symptoms would indicate an alteration in the noradrenergic nervous system is occurring early in the progression of PD. Postmortem examination of PD tissue demonstrates a significant loss of noradrenergic neurons in the locus coeruleus (LC); this loss is equal to or greater than the neuronal loss observed in the SN (Bertrand et al., 1997; Cash et al., 1987; Chan-Palay & Asan, 1989; Hornykiewicz & Kish, 1987; Marien et al., 2004; McMillan et al., 2011; Patt & Gerhard,

**1. Introduction** 

**1.1 Dopaminergic neuronal loss in Parkinson's disease** 

**1.2 Noradrenergic neuronal loss in Parkinson's disease** 

