Project description:Methods: U2OS mRNA profiles of control siRNA(siControl) and siCDK12 were generated by deep sequencing, in triplicate, using Illumina. The sequence reads that passed quality filters were analyzed at the transcript isoform level. Total RNA-Seq was used for normalzation of RIP-Seq signals. Results: We report the application of RNA-immunoprecipitation-based sequencing for high-throughput profiling of mRNA abundence in mammalian cells. We generated genome-wide views of translation rate in CDK12-depleted U2OS cells in comparion to mTOR inhibitor, rapamycin treatment. We find that significant overlap and high corelation of change in eIF4G-associated mRNAs between two condition, rapamycin treatment and siCDK12 transfected cells. This study provides a role of CDK12 in translation control of mammalian cell.
Project description:Methods: U2OS mRNA profiles of control siRNA(siControl) and siCDK12 were generated by deep sequencing, in triplicate, using Illumina. The sequence reads that passed quality filters were analyzed at the transcript isoform level. Total RNA-Seq was used for normalzation of RIP-Seq signals.Results: We report the application of RNA-immunoprecipitation-based sequencing for high-throughput profiling of mRNA abundence in mammalian cells. We generated genome-wide views of translation rate in CDK12-depleted U2OS cells in comparion to mTOR inhibitor, rapamycin treatment. We find that significant overlap and high corelation of change in eIF4G-associated mRNAs between two condition, rapamycin treatment and siCDK12 transfected cells. This study provides a role of CDK12 in translation control of mammalian cell.
Project description:We describe the epigenetic profiling of the H3K9me2 and HP1a in Drosophila third instar larvae before and after CDK12 depletion by RNA interference (RNAi). Here we show that CDK12 regulates heterochromatin dynamics in Drosophila chromosomes. Depletion of CDK12 induces the increased HP1a and H3K9me2 binding profile on the coding region of euchromatic genes, with the X chromosome being the most affected. These results are consistent with the polytene chromosome immunostaining pattern of HP1a and H3K9me2 after CDK12 knockdown in our initial cytological observations, which show that CDK12 depletion induce heterochromatin spreading on euchromatic arms, especially on the X chromosome. This study describes a novel role of the CDK12 complex in controlling the epigenetic transition between euchromatin and heterochromatin. Examination of the genome-wide H3K9me2 and HP1a binding profile in wildtype larvae (WT) and CDK12-depleted larvae (CDK12-KD). Examination of the genome-wide CDK12 binding profile in wildtype larvae (WT). Twelve independent immunoprecipitations were conducted for each antibody. Two biological replicates were performed.
Project description:Determining interaction partners with CDK12 using IP-MS. Capturing global proteome expression changes as a result of CDK12 knockdown.
Project description:In a forward genetic screen, we have previously identified a null mutant of Cdk12 that results in alterations in actin dynamics, the axon initial segment and electrophysiology in Drosophila melanogaster. To decipher how Cdk12 may be having these effects, we extracted RNA from pooled Drosophila heads and compared Cdk12-null mutants to controls at the transcriptome level.