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| Status |
Public on Aug 10, 2016 |
| Title |
Cnot3-dependent mRNA deadenylation safeguards the pluripotent state |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Poly(A)-tail length and mRNA deadenylation play important roles in post-transcriptional gene regulation. Here, we report the regulation of mRNA expression, half-life, and polysome association in mouse embryonic stem cells by the Cnot3 subunit in the Ccr4-Not mRNA deadenylase complex.
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| Overall design |
To determine the consequence of Cnot3 deletion, Cnot3 conditional knockout ESCs were first treated with DMSO (WT) or 0.1μM 4-hydroxytamoxifen (4OHT, KO). 1. In one experiment, cells were collected 3 days after the 4OHT treatment and total RNAs were extracted and sequenced to determine changes in mRNA expression. 2. In the second experiment, cells were collected 3 days after the 4OHT treatment and polysome-associated RNAs were fractionated using a published protocol, purified, and sequenced to determine changes in polysome-association. 3. In the third experiment, cells were further treated with actinomycin D (10ug/ml) 2 days after the initial 4OHT treatment. Cells were collected at 0, 4, 8 hrs after actinomycin D treatment and total RNAs were extracted and sequenced to determine changes in mRNA half-life. Two biological replicates were carried out for each experiment. RNAs were extracted using RNA purification kits from Life Technologies, and RNA-seq libraries were prepared using Illumina TruSeq RNA library preparation kit.
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| Contributor(s) |
Guang H, Zheng X |
| Citation missing |
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| Submission date |
Jul 28, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Guang Hu |
| E-mail(s) |
guang.hu@nih.gov
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| Phone |
(919) 541-4755
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| Organization name |
NIH/NIEHS
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| Department |
ESCBL
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| Lab |
Stem Cell Biology Group
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| Street address |
111 T. W. Alexander dr.
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| City |
Res Triangle Park |
| State/province |
NORTH CAROLINA |
| ZIP/Postal code |
27519 |
| Country |
USA |
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| Platforms (1) |
| GPL13112 |
Illumina HiSeq 2000 (Mus musculus) |
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| Samples (20)
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| Relations |
| BioProject |
PRJNA335741 |
| SRA |
SRP080131 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE84953_RAW.tar |
2.8 Mb |
(http)(custom) |
TAR (of TXT) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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