Proteasome inhibitors suppress formation of polyglutamine-induced nuclear inclusions in cultured postmitotic neurons.
At least nine neurodegenerative disorders are caused by expansion of polyglutamine repeats in various genes. This expansion induces the formation of nuclear inclusions (NI) within various cell types. In this study, we developed a model for polyglutamine diseases using primary cultures of sympathetic neurons from the superior cervical ganglia of prenatal rat pups. Transfection with a plasmid encoding 127 glutamine repeats causes NI to develop in approximately 70% of the sympathetic neurons within 6 days. In addition, it causes somatic atrophy and inhibits dendritic growth. The NIs contain ubiquitinated proteins and sequester the molecular chaperone heat shock protein 70 (Hsp70). We found that two specific proteasome inhibitors, lactacystin and CEP1612, suppress thezformation of polyglutamine-induced NI. In addition, lactacystin treatment induced the removal of preexisting NI. Western blotting and immunocytochemistry revealed that lactacystin and CEP1612 strongly induce the expression of Hsp70, whereas less specific proteasome inhibitor such as N-acetyl-Leu-Leu-Norleucinal does not. Coexpression of 127 glutamines with a plasmid encoding wild-type Hsp70 gene resulted in a marked reduction of the percentage of neurons containing NI. In addition, transfection with plasmids encoding mutant Hsp70 blocked the effects of lactacystin. These findings further implicate Hsp70 as a neuroprotective molecule and they suggest the potential utility of certain proteasome inhibitors in the treatment of polyglutamine diseases.
Kim
Woo-Yang
WY
Department of Pharmacology and Toxicology, School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, New York 14214, USA.
Horbinski
Craig
C
Sigurdson
Wade
W
Higgins
Dennis
D
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D000818:Animals; D015153:Blotting, Western / Q000379:methods; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins / Q000378:metabolism; D002452:Cell Count; D048429:Cell Size; D002478:Cells, Cultured; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D003712:Dendrites / Q000378:metabolism; D004151:Dipeptides / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004622:Embryo, Mammalian; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology; D007150:Immunohistochemistry / Q000379:methods; D045586:Intranuclear Inclusion Bodies / Q000187:drug effects* / Q000502:physiology; D016296:Mutagenesis; D009474:Neurons / Q000187:drug effects* / Q000502:physiology; D010455:Peptides / Q000235:genetics / Q000494:pharmacology* / Q000502:physiology; D010797:Phthalimides / Q000494:pharmacology*; D051381:Rats; D017783:Superior Cervical Ganglion / Q000166:cytology; D013997:Time Factors; D014162:Transfection / Q000379:methods; D016212:Transforming Growth Factor beta / Q000378:metabolism
C525821:Bmp7 protein, rat; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides; D018840:HSP70 Heat-Shock Proteins; D010455:Peptides; D010797:Phthalimides; D016212:Transforming Growth Factor beta; C067713:lactacystin; D049452:Green Fluorescent Proteins; C097188:polyglutamine; C118004:CEP 1612; D000111:Acetylcysteine