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| Status |
Public on Apr 24, 2007 |
| Title |
Chromatin immunoprecipitation (ChIP) assay of CWO protein with Drosophila genome tiling array |
| Organism |
Drosophila melanogaster |
| Experiment type |
Genome binding/occupancy profiling by genome tiling array
|
| Summary |
CWO binding sites were genome-widely searched with Drosophila genome tiling array.
Abstract:
The Drosophila circadian clock consists of integrated autoregulatory feedback loops, making the clock difficult to elucidate without comprehensively identifying the network components in vivo. Previous studies have adopted genome-wide screening for clock-controlled genes using high-density oligonucleotide arrays that identified hundreds of clock-controlled genes. In an attempt to identify the core clock genes among these candidates, we applied genome-wide functional screening using an RNAi system in vivo. Here we report the identification of novel clock gene candidates including clockwork orange (cwo), a transcriptional repressor belonging to the basic helix-loop-helix-ORANGE family. cwo is rhythmically expressed and directly regulated by CLK-CYC through canonical E-box sequences. A genome-wide search for its target genes using the Drosophila genome tilling array revealed that cwo forms its own negative feedback loop and directly suppresses the expression of other clock genes through the E-box sequence. Furthermore, this negative transcriptional feedback loop contributes to sustaining a high-amplitude circadian oscillation in vivo. Based on these results, we propose that the competition between cyclic CLK-CYC activity and the adjustable threshold imposed by CWO keeps E-box-mediated transcription within the controllable range of its activity, thereby rendering a Drosophila circadian clock capable of generating high-amplitude oscillation.
Keywords: ChIP-chip
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| Overall design |
4 Drosophila genome tiling arrays were used as follows:
Array1 Immunoprecipetated with anti-Flag antibody (Biological replicate no.1) for CWO binding signal
Array2 Immunoprecipetated with anti-Flag antibody (Biological replicate no.2) for CWO binding signal
Array3 Immunoprecipetated with anti-V5 antiboy (Biological replicate no.1) for background signal
Array4 Immunoprecipetated with anti-V5 antiboy (Biological replicate no.2) for background signal
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| Contributor(s) |
Matsumoto A, Tadenuma MU, Yamada RG, Houl J, Uno KD, Kasukawa T, Dauwalder B, Itoh TQ, Takahashi K, Ueda R, Hardin PE, Tanimura T, Ueda HR |
| Citation(s) |
17578908 |
| |
| Submission date |
Apr 22, 2007 |
| Last update date |
Jul 24, 2013 |
| Contact name |
Rikuhiro G Yamada |
| E-mail(s) |
rikuhiro@cdb.riken.jp
|
| Phone |
+81-78-306-3191
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| Fax |
+81-78-306-3194
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| Organization name |
RIKEN
|
| Department |
Center for Developmental Biology
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| Lab |
Laboratory for Systems Biology
|
| Street address |
Minatojima-Minamimachi 2-2-3
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| City |
Kobe |
| State/province |
Hyogo |
| ZIP/Postal code |
650-0047 |
| Country |
Japan |
| |
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| Platforms (1) |
| GPL3923 |
Affymetrix Drosophila Melanogaster Antisense Tiling Array |
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| Samples (4)
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| GSM183500 |
ChIP-Chip with anti-Flag antibody, replicate 1 of 2. |
| GSM183501 |
ChIP-Chip with anti-Flag antibody, replicate 2 of 2. |
| GSM183502 |
ChIP-Chip with anti-V5 antibody, replicate 1 of 2. |
| GSM183503 |
ChIP-Chip with anti-V5 antibody, replicate 2 of 2. |
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| Relations |
| BioProject |
PRJNA100339 |
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