scRNAseq analysis of lytically and latently infected KSHV-infected iSLK.219 cells with and without caspase inhibitor treatment

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. iSLK.219 cells were treated with doxycyline to induce lytic reactivation (as per Myoung and Ganem, 2011) or left untreated for latently infected cells. The doxycycline induces expression of an exogenous copy of the KSHV lytic regulator RTA. For caspase inhibitor treated samples, they were treated with the pan-caspase inhibitor IDN-6556 and for anti-inteferon antibody treatment, they were treated with a mixture of monoclonal and polyclonal antibodies directed against human type I Interferons and type I Interferon Receptor Subunit 2 (IFNAR2). Untreated samples received vehicle. All drugs were replenished two days after the start of reactivation and treatment, and samples for scRNAseq were collected 4 days after the start of treatment. Uninfected iSLK.H cells were mixed with vehicle-treated uninduced iSLK.219 cells before single-cell analysis. The 10x Genomics platform was used to measure single-cell RNA levels.