Project description:MDA231, BT549, and SUM159PT basal-like breast cancer cell lines were transfected with non-targeting siRNA (siCONTROL), siRNA targeting DUSP4 (siDUSP4), or siCONTROL + 4 or 24 hr of 1uM selumetinib. Cells were harvested at 96 hr post-siRNA transfection. Data were Log2 RMA normalized.
Project description:MDA231, BT549, and SUM159PT basal-like breast cancer cell lines were transfected with non-targeting siRNA (siCONTROL), siRNA targeting DUSP4 (siDUSP4), or siCONTROL + 4 or 24 hr of 1uM selumetinib. Cells were harvested at 96 hr post-siRNA transfection. Data were Log2 RMA normalized. We sought to identify changes in gene expression after MEK inhibition, or after loss of DUSP4 function in breast cancer cell lines.
Project description:A RNA-seq analysis was performed to characterize the molecular intermediates responsible for the deleterious effects upon DUSP4 loss in SKMEL28 cells. Methods:SKMEL28 cells were transfected with siRNA against DUSP4 or non-targeting control with or without Trametinib (Tram) at 0.25nM. After 16 hours, cell were collected and RNAseq experiments were performed. Results: the expression of MITF and some of its target genes were significantly downregulated in the absence of DUSP4 and completely rescued by trametinib treatment. Conclusions: In mutant melanoma cells, the expression of MITF and its related target genes is subject to regulation by the DUSP4-ERK axis.
Project description:In order to confirm the role of fatty acid β-oxidation in Src regulation, we performed gene expression analysis in MDA231 cells from in vivo model treated with ETX or knockdown of CPT1 or CPT2 using shRNA. As expected, inhibition of β-oxidation showed a gene expression pattern that is opposite to the published Src regulated gene pattern. The known Src up-regulated genes are down-regulated and Src down-regulated genes are up-regulated in β-oxidation inhibited cells. Western Blotting further confirmed the gene expression pattern. Knockdown of CPT1 or CPT2 inhibited Src Y416 autophosphorylation as observed with ETX. MDA231 cells were treated with ETX or knockdown of CPT1 or CPT2 using shRNA. Gene expression profiles were taken for each group and compared with control group (shRNA scramble). Multiple group comparison [MDA231-Scramble (C), MDA231-Scramble +ETX (D), MDA231-shCPT-1 (E) and MDA231-shCPT-2 (F)]
Project description:To further study the regulation of SIPA1 on cells, the transcriptome sequencing on BT549 cells, BT549/shSIPA1 cells and si-dDBR cells were performed. We then performed gene expression profiling analysis using data obtained from RNA-seq of BT549 cells, BT549/shSIPA1 cells and si-dDBR cells to find the genes regulated by SIPA1.
Project description:Purpose: Gene expression analysis of knockdown and overexpression of LBH (Limb-Bud-and-Heart) in human breast cancer cell lines using RNA-Seq Methods: RNA was collected and analyzed from three biological replicates of each condition (LBH vs vector) for LBH overexpression (OE) in two human breast cancer cell lines (BT549, MCF7). Additionally, RNA was collected and analyzed from three biological replicates of each condition (siLBH vs non-target/NT siRNA) for LBH knockdown (KD) in two human triple negative breast cancer cells lines (HCC1395, MDA-MB-231). High-throughput sequencing used Illumina platforms. Results: Using an optimized data analysis workflow, we mapped about 60 million sequence reads per sample to the human genome (build hg19). Conclusions: Our study represents the first gene profiling analysis of LBH transcriptomes in human breast cancer cell lines, with biologic replicates, generated by RNA-seq technology.
Project description:Mitogen-activated protein kinases are inactivated by dual specificity phosphatases (DUSPs), whose activities are tightly regulated during cell differentiation. Using knockdown screening and single-cell transcriptional analysis, we determined that DUSP4 is the phosphatase that specifically inactivates p38 kinase for the promotion of megakaryocyte (Mk) differentiation. Mechanistically, PRMT1-mediated methylation of DUSP4 triggers its ubiquitinylation by an E3 ligase HUWE1. Interestingly, the mechanistic axis of the DUSP4 degradation and p38 activation is also associated with a transcriptional signature of immune activation in Mk cells. In the context of thrombocytopenia observed in myelodysplastic syndromes (MDS), we demonstrated that high levels of p38 MAPK and PRMT1 are associated with low platelet counts and adverse prognosis, while pharmacological inhibition of p38 MAPK or PRMT1 stimulates megakaryopoiesis. These findings provide mechanistic insights into the role of the PRMT1-DUSP4-p38 axis on Mk differentiation and present a targeting strategy for treatment of thrombocytopenia associated with MDS.
