title: Novel mechanism of Hsp70 chaperone-mediated prevention of polyglutamine aggregates in a cellular model of huntington disease.
abstract: The key feature of polyglutamine aggregates accumulating in the course of Huntington disease (HD) is their resistance to protein denaturants, and to date only chaperones are proved to prevent mutant protein aggregation. It was suggested that expanded polyglutamine chains (polyQ) of mutant huntingtin are cross-linked to other proteins such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Here we clarify the roles of GAPDH and molecular chaperone Hsp70 in the formation of sodium dodecyl sulfate (SDS)-insoluble polyQ aggregates. First, the addition of pure GAPDH was found to enhance the aggregation of polyQ in a cell-free model of HD. Secondly, the immunodepletion of GAPDH dose-dependently decreased polyQ aggregation. Finally, siRNA-mediated inhibition of GAPDH protein in SK-N-SH neuroblastoma cells has also reduced the aggregation of cellular polyQ. Regulated over-expression of Hsp70 decreased the amount of GAPDH associated with SDS-insoluble polyQ aggregates. Physical association of Hsp70 and GAPDH in SK-N-SH cells was shown by reciprocal immunoprecipitation and confocal microscopy. Pure Hsp70 dose-dependently inhibited the formation of polyQ aggregates in cell-free model of HD by sequestering both GAPDH and polyQ. We demonstrated that Hsp70 binds to polyQ in adenosine triphosphate-dependent manner, which suggests that Hsp70 exerts a chaperoning activity in the course of this interaction. Binding of Hsp70 to GAPDH was nicotinamide adenine dinucleotide-dependent suggesting another type of association. Based on our findings, we conclude that Hsp70 protects cells in HD by removing/sequestering two intrinsic components of protein aggregates: the polyQ itself and GAPDH. We propose that GAPDH might be an important target for pharmacological treatment of HD and other polyglutamine expansion-related diseases.
authors.affiliation: Institute of Cytology of Russian Academy of Sciences, Tikhoretsky pr., 4, 194064 St Petersburg, Russia. guzhova@mail.cytspb.rssi.ru
authors.forename: Irina V
authors.initials: IV
authors.lastname: Guzhova
authors.forename: Vladimir F
authors.initials: VF
authors.lastname: Lazarev
authors.forename: Anastasia V
authors.initials: AV
authors.lastname: Kaznacheeva
authors.forename: Maria V
authors.initials: MV
authors.lastname: Ippolitova
authors.forename: Vladimir I
authors.initials: VI
authors.lastname: Muronetz
authors.forename: Alexander V
authors.initials: AV
authors.lastname: Kinev
authors.forename: Boris A
authors.initials: BA
authors.lastname: Margulis
mesh_terms: D045744:Cell Line, Tumor; D015870:Gene Expression; D005786:Gene Expression Regulation; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006816:Huntington Disease / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism*; D011485:Protein Binding; D012995:Solubility
chemical_list: D018840:HSP70 Heat-Shock Proteins; D010455:Peptides; C097188:polyglutamine; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases