ABSTRACT: To study the senescence gene signatures in the cells, which were genetic SMARCB1 depleted or treated with aurora kinase inhibitors or etoposide, we performed next generation RNA sequencing on these cell, and 'FRIDMAN_SENESCENCE_UP' geneset was used to determine the enrichment of senescence-related genes. The RNA sequencing results include (1) A375 cells and SMARCB1 depleted counterparts. (2) A549 cells and aurora kinase inhibitor (Alisertib, barasertib or tozasertib) or etoposide treated counterparts.
Project description:We analyzed a panel of 13 cancer cell lines rendered senescent by either etoposide (7 day treatment) or alisertib (7 and 14 day treatment). We aimed to find common vulnerabilities to induce cell death, and shared features that would allow unambiguous identification of the senescent state in the context of a cancerous phenotype. The samples were treated for the given duration, followed by 24 hours without drug before RNA was extracted. The senescence phenotype was assessed by B-gal staining, growth arrest and morphology.
Project description:Determine the effect of knocking out the H2afj gene on the transcriptome of MEFs induced into senescence with etoposide. MEFs were produced from individual H2A.J-ko or isogenic WT C57BL/6-N embryos. 3 different female WT and H2A.J-ko MEFs were used for the transcriptome analyses. MEFs were cultivated in DMEM + Gluta-Max (Gibco 31966) + 10%FBS + pen/strep in a 5% carbon dioxyde and 5% oxygen incubator. MEFs were passed twice before inducing senescence by treatment with 2.5 µM etoposide for 6 days (with media changed after 3 days), followed by 3 days incubation in fresh medium without etoposide.
Project description:Study the role of H2A.J in gene expression in senescent human fibroblasts by comparing RNA-seq of WI-38hTERT fibroblasts expressing either an sh2-H2AFJ or sh-NoTarget RNAs. Senescence was induced by treating cells with 20 M etoposide for 2 weeks followed by one further week of incubation without etoposide. 3 biological replicates of sh2-H2AFJ and sh-NoTarget were sequenced.
Project description:Aurora-A has attracted a great deal of interest as a potential therapeutic target. However, the outcomes of inhibitors targeting Aurora-A are not as favorable as expected, and the basis of their ineffectiveness remains unknown. Here, we found that signal transducer and activator of transcription 1 (STAT1) was highly expressed in colorectal cancer (CRC) xenograft mouse models that were resistant to alisertib, an Aurora-A inhibitor, in an interferon/Janus kinase-independent manner, suggesting an unconventional mechanism regulating STAT1 expression. Unexpectedly, we found that alisertib disrupted Aurora-A binding with ubiquitin-like with plant homeodomain and ring finger domain 1 (UHRF1), leading to UHRF1-mediated ubiquitination and degradation of DNA methyltransferase 1 (DNMT1), which in turn resulted in demethylation of the CpG islands in the STAT1 promoter and STAT1 overexpression. Simultaneous silencing of Aurora-A and UHRF1 prevented STAT1 overexpression and effectively inhibited CRC growth. Hence, concomitant targeting of Aurora-A and UHRF1 can be a promising therapeutic strategy for cancer patients.
Project description:The intent of the experiment was to identify differentially expressed micro-RNAs comparing WI-38hTERT/sh3-H2AFJ and WI-38hTERT/sh-NT fibroblasts in proliferation and etoposide-induced senescence.
Project description:The intent of the experiment was to compare H3-K4me3 peaks on the chromatin of WI-38hTERT/sh3-H2AFJ and WI-38hTERT/sh-NT fibroblasts in proliferation and etoposide-induced senescence.
Project description:The intent of the experiment was to identify differentially expressed RNAs comparing WI-38hTERT/sh3-H2AFJ and WI-38hTERT/sh-NT fibroblasts in proliferation and etoposide-induced senescence.
Project description:The intent of the experiment was to compare RNA polymerase II occupancy on the chromatin of WI-38hTERT/sh3-H2AFJ and WI-38hTERT/sh-NT fibroblasts in proliferation and etoposide-induced senescence.
Project description:The Aurora-A inhibitor alisertib shows encouraging activities in clinical trials against multiple malignances including advanced breast cancer. However, its mechanism of action remains unclear, especially regarding how the inflammatory microenvironment is involved in the efficacy of alisertib. Here, we demonstrated that Aurora-A inhibition directly reshapes the immune microenvironment through removal of tumor-promoting myeloid cells and enrichment of anti-cancer T lymphocytes, which restores a tumor-suppressive microenvironment and significantly contributes to the regression of murine mammary tumors. Mechanistically, the Aurora-A inhibitor effectively eliminated myeloid cells including myeloid-derived suppressor cells (MDSCs) and macrophages in tumors by triggering apoptosis of these cells. Further, Aurora-A inhibition could disrupt the immunosuppressive functions of MDSCs through inhibiting Stat3 mediated ROS production. These alterations led to significant increases in the proportion and the number of CD8+ and CD4+ T lymphocytes, which efficiently inhibited the proliferation of tumor cells. In summary, these data revealed that in addition to suppressing the proliferation of tumor cells, Aurora-A inhibitor directly modulates and restores an anti-tumor immune-microenvironment in breast cancer. Intriguingly, Aurora-A inactivation combined with PD-L1 blockade showed synergistic efficacy in the treatment of mammary tumors, providing an effective strategy for clinical trials of chemo-immunotherapy in breast cancer.
Project description:BCLAF1 is a serine-arginine (SR) protein implicated in transcriptional regulation and mRNA splicing. We have recently identified BCLAF1 as part of a novel mRNA splicing complex that is recruited to different genetic promoters by the breast cancer susceptiblity protein, BRCA1 in response to DNA damage. This ChIP-chip study was designed to identify genes/promoters regulated by the BRCA1/BCLAG1 mRNA splicing complex by identifying promoters bound by BCLAF1 in the absense and presense of BRCA1 in control cells and cells treated with etoposide to induce DNA damage. This study includes tripicate BCLAF1 ChIP-chip experiments in untreated and etoposide treated (1uM 16 hours) control cells (siGFP) and cells depleted of BRCA1 (siBRCA1).