Project description:Expression profiling of stably infected epithelial cells using a custom tiling microarray. iSLK was infected with rKSHV.219 and selected with puromycin. Mock infected iSLK served as control for iSLK.219. Lytic reactivation of iSLK.219 was induced with 1 ug/mL doxycycline for 48 hours.
Project description:Expression profiling of stably infected primary endothelial cells using a custom tiling microarray. LEC and BEC were infected with rKSHV.219 and selected with puromycin for 2 weeks. Mock infected LEC and BEC served as controls for their infected counterparts. All cells were harvested after 2 weeks of stable infection.
Project description:We used ac4C-seq to detedc cellular and KSHV transcripts in iSLK-Puro, iSLK-KSHV, iSLK-KSHV (WT), iSLK-KSHV (ΔNAT10) cells after after doxycycline and sodium butyrate treatment.
Project description:The goal of this analysis was to examine the cell-to-cell variation in Kaposi's sarcoma-associated herpesvirus (KSHV) reactivation and type I interferon response after caspase inhibitor treatment. We profiled the levels of host and viral mRNAs at a single-cell level using the iSLK.219 tissue culture model system (Myoung and Ganem, 2011). We examined gene expression in lytically reactivating cells, lytically reactivating cells treated with caspase inhibitors, and lytically reactivating cells treated with casapse inhibitors and neutralizing antibodies against interferons. (we previously described that caspase inhibitors elicit an interferion response in KSHV-infected cells, Tabtieng et al., 2018). We also examined gene expression in a mixture of latently infected cells and uninfected cells as a control.
Project description:Expression profiling of stably infected primary endothelial cells using a custom tiling microarray. LEC, BEC, HUVEC, and HAEC were infected with rKSHV.219 and selected with puromycin for 2 weeks. Mock infected LEC, BEC, HUVEC, and HAEC served as controls for their infected counterparts. All cells were harvested after 2 weeks of stable infection.
Project description:To investigate the regulation of ER stress-related gene expression by KSHV-ORF45 during lytic replication, we performed RNA-sequencing analysis of iSLK-BAC16 vs. iSLK-STOP45 cells under lytic induction for 72h. When the differentially expressed genes were filtered and analyzed, we found that ER stress-related gene expression was much low in iSLK-STOP45 cells compared with iSLK-BAC16 cells, indicating that ORF45 expression is required for induction of ER stress.To further reveal the signal transduction of LAMP3 in KSHV lytic replication, RNA-sequencing analysis was performed to identify the differentially expressed genes (DEGs) in normal vs. LAMP3-silenced cells under lytic replication. A total of 35372 raw read targets were obtained and all DEG clusters were filtered and analyzed by KEGG pathway enrichment analysis. Eleven pathway were significantly enriched over 10 fold in LAMP3-depleted cells compared with control cells, including PI3K-Akt signaling pathway.Given that Akt and ERK activation play the important roles in KSHV lytic replication, we conclude that LAMP3 might promote Akt and ERK activation and then consequently facilitate KSHV lytic replication.
Project description:Methylation at the N6 position of adenosine (m6A) is a highly prevalent reversible modification within eukaryotic mRNAs that has been linked to many stages of RNA processing and fate. Recent studies suggest that m6A deposition and proteins involved in the m6A pathway play a diverse set of roles in either restricting or modulating the lifecycles of select viruses. Here, we report that m6A levels are significantly increased in cells infected with the oncogenic human DNA virus Kaposi’s sarcoma-associated herpesvirus (KSHV). Transcriptome-wide m6A-sequencing of the KSHV-positive renal carcinoma cell line iSLK.219 during lytic reactivation revealed the presence of m6A across multiple kinetic classes of viral transcripts, and a concomitant decrease in m6A levels across much of the host transcriptome.
Project description:Herpes simplex virus 1 (HSV-1) switches between lytic and latent infections in neurons, yet the switch mechanisms remain obscure. Here we identify forkhead box (FOX) family proteins that can strongly activate or repress HSV-1 replication particularly in neuronal cells. Expression of activating FOX genes (Foxa, Foxc, Foxe, Foxf) is ordinarily low and expression of repressive FOX genes (Foxk) is high in neurons. Lytic HSV-1 infection and other stresses can increase activating FOX gene expression. Such overexpression or knockout of endogenous FOXK1 promotes reactivation from latency. These FOX proteins broadly associate with the viral genome and regulate viral gene transcription through epigenetic modulators. FOXF1 associates with histone acetyltransferases CBP and P300 to open viral chromatin. FOXK1 collaborates with SIN3A, a known cofactor of histone deacetylation, and MAX to suppress HSV-1 replication and antagonize activating FOX proteins. Thus, HSV-1 uses FOX family members with opposing effects to regulate the lytic-latent balance.
