The identification of chemical compounds that decrease cellular levels of toxic Huntington's disease protein through a novel cell-based assay

Huntington's disease (HD) is a progressive degenerative neurological disorder. Individuals who inherit the IT15 gene with an expansion of the CAG repeat region inevitably succumb to increasingly sever motor, psychological, and cognitive symptoms. I sought to develop an assay system with the capability for identification of chemical compounds that selectively decrease the intracellular levels of disease-causing expanded polyglutamine huntingtin (Htt) protein without reducing the intracellular levels of the potentially protective normal Htt. To achieve this goal I designed a cell-based assay using the enzymatic activity of E. coli [beta]-galactosidase as a reporter for Htt protein levels. I expressed either expanded (97Q) or normal (23Q) Htt fused to the [beta]-galactosidase alpha-subunit ([alpha]) in an inducible fashion in PC12 cells which also expressed the [beta]-galactosidase delta-subunit ([delta]). Complementation between these expressed subunits allowed the formation of functional P-galactosidase. The level of [beta]-galactosidase activity in these [delta]-[alpha]97Q and [delta]-[alpha]23Q cells directly correlated with the amount of a 97Q and a 23Q fusion protein levels, indicating that [beta]-galactosidase activity could be used as a reporter in this system for Htt protein levels.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Biology.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2006

Author dc:creator

• Coufal, Myra Alfert

Advisor dc:contributor.advisor

• David E. Housman.

• Biology.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/34622
• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Identifier URI

http://dspace.mit.edu/handle/1721.1/34622

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/34622