PT  - JOURNAL ARTICLE
AU  - Kenichi Mitsui
AU  - Hiroshi Nakayama
AU  - Takumi Akagi
AU  - Munenori Nekooki
AU  - Kenji Ohtawa
AU  - Koji Takio
AU  - Tsutomu Hashikawa
AU  - Nobuyuki Nukina
TI  - Purification of Polyglutamine Aggregates and Identification of Elongation Factor-1α and Heat Shock Protein 84 as Aggregate-Interacting Proteins
AID  - 10.1523/JNEUROSCI.22-21-09267.2002
DP  - 2002 Nov 01
TA  - The Journal of Neuroscience
PG  - 9267--9277
VI  - 22
IP  - 21
4099  - http://www.jneurosci.org/content/22/21/9267.short
4100  - http://www.jneurosci.org/content/22/21/9267.full
SO  - J. Neurosci.2002 Nov 01; 22
AB  - Aggregates of green fluorescent protein (GFP)-fused truncated N-terminal huntingtin containing abnormally long polyglutamine tracts (150 repeats of glutamine residue) were purified from an ecdysone-inducible mutant neuro2A cell line (HD150Q-28) by using a fluorescence-activated cell sorter. To analyze the aggregate-interacting proteins, we subjected the purified aggregates to SDS-PAGE; prominent protein bands in the gel were digested with Achromobactor lysyl endopeptidase, followed by a HPLC-mass spectrometry (MS) analysis. The resulting data of tandem MS analysis revealed that, in addition to ubiquitin and widely reported chaperone proteins such as heat shock cognate 70 (HSC70), human DNA J-1 (HDJ-1), and HDJ-2, the translational elongation factor-1α (EF-1α) and heat shock protein 84 (HSP84) also were recognized as aggregate-interacting proteins. Sequestration of these proteins to aggregates was confirmed further by several immunochemical methods. We confirmed that, in addition to the other known proteins, EF-1α and HSP84 also colocalized with the intracellular aggregates. An assay of the transient expression of EF-1α and HSP84 in HD150Q-28 cells revealed that both proteins improved cell viability. Moreover, the rate of aggregate formation decreased in both transfectants. Our study suggests that both EF-1α and HSP84 are involved in the neurodegenerative process of polyglutamine diseases.