title: D-polyglutamine amyloid recruits L-polyglutamine monomers and kills cells.
abstract: Polyglutamine (polyQ) amyloid fibrils are observed in disease tissue and have been implicated as toxic agents responsible for neurodegeneration in expanded CAG repeat diseases such as Huntington's disease. Despite intensive efforts, the mechanism of amyloid toxicity remains unknown. As a novel approach to probing polyQ toxicity, we investigate here how some cellular and physical properties of polyQ amyloid vary with the chirality of the glutamine residues in the polyQ. We challenged PC12 cells with small amyloid fibrils composed of either L- or D-polyQ peptides and found that D-fibrils are as cytotoxic as L-fibrils. We also found using fluorescence microscopy that both aggregates effectively seed the aggregation of cell-produced L-polyQ proteins, suggesting a surprising lack of stereochemical restriction in seeded elongation of polyQ amyloid. To investigate this effect further, we studied chemically synthesized D- and L-polyQ in vitro. We found that, as expected, D-polyQ monomers are not recognized by proteins that recognize L-polyQ monomers. However, amyloid fibrils prepared from D-polyQ peptides can efficiently seed the aggregation of L-polyQ monomers in vitro, and vice versa. This result is consistent with our cell results on polyQ recruitment but is inconsistent with previous literature reports on the chiral specificity of amyloid seeding. This chiral cross-seeding can be rationalized by a model for seeded elongation featuring a "rippled β-sheet" interface between seed fibril and docked monomers of opposite chirality. The lack of chiral discrimination in polyQ amyloid cytotoxicity is consistent with several toxicity mechanisms, including recruitment of cellular polyQ proteins.
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA; Pittsburgh Institute for Neurodegenerative Diseases, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA.
authors.forename: Karunakar
authors.initials: K
authors.lastname: Kar
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA; Pittsburgh Institute for Neurodegenerative Diseases, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA.
authors.forename: Irene
authors.initials: I
authors.lastname: Arduini
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA; Pittsburgh Institute for Neurodegenerative Diseases, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA.
authors.forename: Kenneth W
authors.initials: KW
authors.lastname: Drombosky
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA. Electronic address: pvdwel@pitt.edu.
authors.forename: Patrick C A
authors.initials: PC
authors.lastname: van der Wel
authors.affiliation: Department of Structural Biology, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA; Pittsburgh Institute for Neurodegenerative Diseases, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA. Electronic address: rwetzel@pitt.edu.
authors.forename: Ronald
authors.initials: R
authors.lastname: Wetzel
mesh_terms: D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D006816:Huntington Disease / Q000378:metabolism; D007536:Isomerism; D007700:Kinetics; D008969:Molecular Sequence Data; D016716:PC12 Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D010455:Peptides / Q000138:chemical synthesis / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D011487:Protein Conformation; D051381:Rats; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D000682:Amyloid; D010455:Peptides; C097188:polyglutamine
keywords: DLS; EM; FTIR; Fourier transform infrared; HD; Huntington's disease; PBS; chiral cross-seeding; dynamic light scattering; electron microscopy; phosphate-buffered saline; recruitment; rippled β-sheet; toxicity