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| Status |
Public on Nov 22, 2011 |
| Title |
Gene and pathways affected by CAG-repeat RNA-based toxicity in Drosophila |
| Organism |
Drosophila melanogaster |
| Experiment type |
Expression profiling by array
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| Summary |
Spinocerebellar ataxia type 3 (SCA3) is one of the polyglutamine (polyQ) diseases, which are caused by a CAG repeat expansion within the coding region of the associated genes.
The CAG repeat specifies glutamine, and the expanded polyQ domain with mutation confers dominant toxicity on the protein.
Traditionally, studies have focused on protein toxicity in polyQ disease mechanisms.
Recent findings, however, demonstrate that the CAG repeat RNA, which encodes the toxic polyQ protein, also contributes to the disease in Drosophila.
To provide insight into the nature of the RNA toxicity,
we extracted brain-enriched RNA from flies expressing a toxic CAG repeat mRNA (CAG100) and a non-toxic interrupted CAA/G mRNA repeat (CAA/G105) for microarray analysis.
This approach identified a set of genes that are differentially expressed specifically in CAG100 flies.
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| Overall design |
Four independent replicates of flies expressing CAG0, CAG100, or CAA/G105 by elav-GAL4 were collected at 3 days.
The transgenes are DsRed with either (CAG0) no CAG repeat in the 3'UTR, (CAG100) a CAG repeat of 100 CAGs in the 3'UTR, or (CAA/G105) an interrupted CAA CAG repeat in the 3'UTR (ref: Li et al., Nature 453:1107)
The transgenes were adjusted to match in mRNA expression such that CAG0 flies had one copy of the transgene, CAG100 flies had 5 copies, and CAA/G105 had two copies.
Fly brain tissue (about 20 brains per sample, dissected from head capsule, eyes, lamina and outer medulla removed) was dissected in cold phosphate buffered saline (PBS) and stored in Trizol reagent (Invitrogen Corporation, Carlsbad, CA) at -80˚C.
Total brain RNA was extracted and purified using TRIzol reagent (Invitrogen) and the RNeasy Mini system (Qiagen), and treated with RNase-free DNase I (Qiagen).
To define genes whose expression is altered in response to a toxic CAG repeat RNA, we compared CAG100 flies with age-matched flies expressing CAG0.
To exclude transcriptional changes in response to a non-toxic trinucleotide repeat, a second gene list was generated by comparing CAA/G105 flies with age-matched CAG0 flies.
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| Contributor(s) |
Shieh S, Bonini NM |
| Citation(s) |
21933837 |
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| Submission date |
Sep 05, 2011 |
| Last update date |
Aug 28, 2018 |
| Contact name |
Bonini Nancy |
| Organization name |
University of Pennsylvania
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| Street address |
415 S University Ave
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| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19104 |
| Country |
USA |
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| Platforms (1) |
| GPL1322 |
[Drosophila_2] Affymetrix Drosophila Genome 2.0 Array |
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| Samples (12)
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| Relations |
| BioProject |
PRJNA145239 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE31875_RAW.tar |
25.7 Mb |
(http)(custom) |
TAR (of CEL) |
| Processed data included within Sample table |
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