Rapid aggregate formation of the huntingtin N-terminal fragment carrying an expanded polyglutamine tract.

Huntington's disease (HD) is caused by an expansion of the CAG repeat in the HD gene. The repeat is translated to the polyglutamine tract as huntingtin, the product of HD gene. Several studies showed that the expansion of polyglutamine tract leads to formation of cytoplasminc and/or intranuclear aggregates in vivo or in vitro. To understand the molecular mechanism of the aggregate formation, we studied the transient expression of HD exon 1-GFP fusion proteins in COS-7 cells. The fusion protein carrying 77 glutamine repeats aggregated in a time-dependent manner, while the fusion protein carrying 25 glutamine tract remained to be distributed diffusely in the cytoplasm even 72 hours after transfection. Initially, fluorescent signals were diffusely distributed in the COS-7 cells that were transfected with the construct containing the 77 CAG repeats. Approximately 40 hours later after the transfection, large aggregates grew very rapidly in those cells and the diffuse cytoplasmic fluorescence faded out. This process was completed within 40 minutes from the appearance of small aggregates in the perinuclear regions. The addition of cycloheximide reduced the frequencies of aggregate formation. A possibility was discussed that the aggregate formation was via nucleation. The focal concentration of mutated proteins in neurons may trigger the aggregate formation.