Project description:Ubiquitin Specific Peptidase 7 (USP7) is a deubiquitinase of several important regulatory proteins, and important regulator of TGFb pathway. To investigate their role in cell signaling, we analyzed global mRNA levels in HEK293T cells that were knocked down with shRNAs against USP7 and non-targeting control (CTRL).
Project description:ChIP-seq experiments were performed in normal or USP7 knockdown A375 cells with specific antibodies against USP7, EZH2, H3K27me3 and H2BK120ub1. Consistent with our expectations, the analysis of ChIP-seq data showed that USP7 loss-of-function led to obviously reduction in USP7, EZH2 and H3K27me3 around the USP7-specific peaks, but an upregulation in H2BK120ub1. This study gave us a new understanding that USP7 is required for EZH2 loading and H3K27 trimethylation.
Project description:Purpose: Study of transcriptomic changes upon depletion of USP7 Methods: Melanoma PDX cells were transduced with lentiviral vectors expressing Scrambled(shSCR) or USP7 targeting (shUSP7) shRNA
Project description:USP7, a dominant DUB activity in 3T3-L1 adipocytes and in mouse adipose tissue, increases Tip60 protein levels, and deubiquitinates Tip60 both in intact cells and in vitro. Treatment with a pan deubiquitinase (DUB) inhibitor, or knockdown of USP7, decreases adipogenesis. Transcriptome analysis reveals a common set of cell cycle genes to be co-regulated by both Tip60 and USP7. Knock down of either factor results in impaired mitotic clonal expansion, an early step in adipogenesis. These results therefore reveal deubiquitination of a transcriptional coregulator to be a key mechanism in the regulation of early adipogenesis Mature 3T3-L1 adipocytes were subjected to RNAi-mediated knock down with control(C), USP7(U)- or Tip60(T)-specific oligonucleotides. For this, 4 replicates of differentiated 3T3-L1 cells were transfected with Amaxa technology. Two days after transfection cells were washed twice with PBS twice and lysed in 0.5ml Trizol (Invitrogen). mRNA expression of Tip60 and USP7 was assessed by qRT-PCR. Amplified cRNA samples were labeled with either cy3 or cy5 and put on microarray together with and oppositely labeled common reference sample consisting of cRNA derived from undifferentiated 3T3-L1 cells.
Project description:To investigate the efficacy of CRISPR-Csm complexes for RNA- knockdown in eukaryotes, we quantified transcript abundance in HEK293T cells after targeting several nuclear or cytoplasmic RNAs. RNA-seq demonstrates efficient and specific knockdown of CRISPR-Csm compared to Cas13 and shRNA knockdown.
Project description:We evaluated the role of Arkadia and ESRP2 in HEK293T cells Expression of mRNA in HEK293T cells under the knockdown of Arkadia or ESRP2
Project description:The goal of the study is to identify differentially expressed isoforms in response to SRSF1 knockdown and/or ionizing radiation in HEK293T cells