Project description:RNA viruses are among the most prevalent pathogens and represent a major burden on society. While RNA viruses have been studied extensively, little is known about the processes that occur during the first several hours of infection due to a lack of sensitive assays. Here, we develop a single-molecule imaging assay, virus infection real-time imaging (VIRIM), to study translation and replication of individual RNA viruses in live cells. VIRIM uncovered a striking heterogeneity in replication dynamics between cells and revealed extensive coordination between translation and replication of single viral RNAs. Furthermore, using VIRIM we identify the replication step of the incoming viral RNA as a major bottleneck of successful infection, and identify host genes that are responsible for inhibition of early virus replication. Single-molecule imaging of virus infection represents a powerful tool to study virus replication and virus-host interactions that may be broadly applicable to RNA viruses.
Project description:CRISPR screen: U2OS or U2OS p53KO cells expressing Cas9 were transduced with a whole-genome library of CRISPR sgRNAs, then treated with either DMSO or etoposide. Differential sgRNA abundances were calculated for each condition and used to determine the effect of each single gene knockout on fitness and the drug-induced death rate. RNA-Seq: U2OS or U2OS p53KO cells were cultured with either DMSO or etoposide for 48 hours, and then U2OS cells were incubated in this conditioned media for 8 hours. RNA was collected and use to observe differential expression changes between conditions.
Project description:We used RNA-seq to examine the effect of Myc activation on U2OS cells transcriptome. We also examined these effects in the presence of Torin-1, an inhibitor of mTOR We measure gene expression profiles in U2OS cells containing an inducible Myc expression vector that were induced or mock-treated in duplicates for 36 hours. In addition, we repeated the experiments in the presence of Torin-1, an inhibitor of mTOR.
Project description:We performed ChIP-seq targeting the glucocorticoid receptor (GR) in the U2OS-GR cell line and the androgen receptor (AR) in the U2OS-AR cell line. The cell lines are derived from U2OS ATTC:HTB-96 and stably transfected with an expression construct for either rat GR or human AR, respectively. The U2OS-GR cells were treated with dexamethasone (1 µM) for 90 minutes. The U2OS-AR cells were treated with R1881 (5 nM) for 4 hours.
Project description:We performed ChIP-seq targeting the H3K27ac, H3K4me1, H3K27me3 and H3K9me3 in the U2OS-GR and U2OS-AR cell lines. The cell lines are derived from U2OS ATTC:HTB-96 and stably transfected with an expression construct for either rat GR or human AR, respectively. The U2OS-GR cells were treated with dexamethasone (1 µM) or vehicle (ethanol) for 90 minutes. The U2OS-AR cells were treated with R1881 (5 nM) or vehicle (DMSO) for 4 hours.