title: Formation of morphologically similar globular aggregates from diverse aggregation-prone proteins in mammalian cells.
abstract: Huntington's disease is a progressive neurodegenerative disorder caused by a polyglutamine repeat expansion in the first exon of the huntingtin (Htt) protein. N-terminal Htt peptides with polyglutamine tracts in the pathological range (51-122 glutamines) form high-molecular-weight protein aggregates with fibrillar morphology in vitro, and they form discrete inclusion bodies in a cell-culture model. However, in some studies, formation of discrete Htt inclusions does not correlate well with cell death. We coexpressed N-terminal Htt fragments containing 91 glutamines fused to different affinity tags in HEK293 cells, and we isolated small aggregates by double sequential-affinity chromatography to assure the isolation of multimeric molecules. Transmission electron microscopy and atomic force microscopy revealed the isolated aggregates as globules or clusters of globules 4-50 nm in diameter without any detectable fibrillar species. Because small nonfibrillar oligomers, not mature fibrils, recently have been suggested to be the principal cytotoxic species in neurodegenerative disease, these Htt globular aggregates formed in cells may represent the pathogenic form of mutant Htt.
authors.affiliation: Biosignal Research Center and Graduate School of Science and Technology, Kobe University, 1-1 Rokkodai-cho, Nada, Kobe 657-8501, Japan.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Mukai
authors.forename: Takayuki
authors.initials: T
authors.lastname: Isagawa
authors.forename: Emiko
authors.initials: E
authors.lastname: Goyama
authors.forename: Shuhei
authors.initials: S
authors.lastname: Tanaka
authors.forename: Neil F
authors.initials: NF
authors.lastname: Bence
authors.forename: Atsuo
authors.initials: A
authors.lastname: Tamura
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Ono
authors.forename: Ron R
authors.initials: RR
authors.lastname: Kopito
mesh_terms: D000345:Affinity Labels; D002460:Cell Line; D002846:Chromatography, Affinity; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics; D006801:Humans; D000071058:Huntingtin Protein; D018625:Microscopy, Atomic Force; D008854:Microscopy, Electron; D046912:Multiprotein Complexes; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism / Q000648:ultrastructure; D009687:Nuclear Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism / Q000648:ultrastructure; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D014162:Transfection
chemical_list: D000345:Affinity Labels; C505032:CFTR protein, human; C086055:HTT protein, human; D000071058:Huntingtin Protein; D046912:Multiprotein Complexes; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D049452:Green Fluorescent Proteins