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Series GSE14215 Query DataSets for GSE14215
Status Public on Dec 25, 2008
Title Genome wide identification of targets of the drosha-pasha/DGCR8 complex
Organism Drosophila melanogaster
Experiment type Expression profiling by genome tiling array
Genome binding/occupancy profiling by genome tiling array
Summary Drosha is a type III RNAse, which plays a critical role in miRNA biogenesis. Drosha and its double-stranded RNA-binding partner protein Pasha/DGCR8 likely recognize and cleave miRNA precursor RNAs or pri-miRNA hairpins co-transcriptionally. To identify RNAs processed by Drosha, we used tiling microarrays to examine transcripts after depletion of drosha mRNA with dsRNA in Drosophila Schneider S2 cells. This strategy identified 137 Drosha-regulated RNAs, including 11 putative pri-miRNAs comprising 15 annotated miRNAs. Most of the identified pri-miRNAs seem extremely large, >10 kilobases as revealed by both the Drosha knock down strategy and by RNA PolII chromatin IP followed by Drosophila tiling microarrays. Surprisingly, more than a hundred additional RNAs not annotated as miRNAs are under Drosha control and are likely to be direct targets of Drosha action. This is because many of them encode annotated genes, and unlike bona fide pri-miRNAs, they are not affected by depletion of the miRNA processing factor, dicer-1. Moreover, application of the evofold analysis software indicates that at least 25 of the Drosha-regulated RNAs contain evolutionarily conserved hairpins similar to those recognized by the Drosha-Pasha/DGCR8 complex in pri-miRNAs. One of these hairpins is located in the 5′ UTR of both pasha and mammalian DGCR8. These observations suggest that a negative feedback loop acting on pasha mRNA may regulate the miRNA-biogenesis pathway: i.e., excess Drosha cleaves pasha/DGCR8 primary transcripts and leads to a reduction in pasha/DGCR8 mRNA levels and Pasha/DGCR8 synthesis.

Keywords: time course, ChIP-chip
Overall design S2 cells were depleted of drosha or dicer mRNA with dsRNA. Total RNA was then converted to cDNA and hybridized to Affymetrix tiling arrays to identify pri-miRNAs and drosha regulated RNAs. RNA PolII chromatin IP genomic DNA was also hybridized to Affymetix tiling arrays to support pri-miRNA targets from drosha knockdowns. Two replicates were used for each sample.
Contributor(s) Kadener S, Rodriguez J, Abruzzi K, Khodor YL, Sugino K, Marr M, Nelson S, Rosbash M
Citation(s) 19223442
Submission date Dec 24, 2008
Last update date Jul 08, 2015
Contact name Joseph Rodriguez
Organization name Brandeis University
Department Biology
Lab Rosbash
Street address 415 South Street
City Waltham
ZIP/Postal code 02453
Country USA
Platforms (2)
GPL5919 [Dm35b_MR] Affymetrix Drosophila Tiling 1.0R Array
GPL6629 [DM_tiling2_MR] Affymetrix Drosophila Tiling 2.0R Array
Samples (7)
GSM356322 S2 control,biological replicate 1
GSM356323 S2 control,biological replicate 2
GSM356324 S2 dicer knockdown,biological replicate 1
BioProject PRJNA111341

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE14215_RAW.tar 373.2 Mb (http)(custom) TAR (of BAR, CEL)
Processed data provided as supplementary file

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