CD8+ T cell responsiveness to anti-PD-1 is epigenetically regulated by Suv39h1 in melanomas [ATAC-seq]

Purpose: We show that genetic ablation or pharmacological inhibition of H3K9-methyl transferase Suv39h1 delays tumor growth and potentiates tumor rejection by anti-PD-1. Suv39h1-KO and littermate WT mice were injected with B16-OVA tumor cells. On day 20, tumors were processed with Liberase and DNase I, stained with DAPI, CD45.2, TCRb, CD8, CD19, NK1.1, F4/80 and sorted on an Aria (BD) by gating on live, CD45.2+, (CD19, NK1.1, F4/80 negative cells) and CD8+ on PBS 0.04% BSA. Chromatin profiling was performed by ATAC-seq as described in Buenrostro et al. In brief, 50,000 cells per sample were washed in cold PBS and lysed. Transposition reaction was performed at 37°C for 30 minutes and DNA was purified with MinElute PCR purification kit (Qiagen). Transposed DNA was amplified for 5 cycles and additional PCR cycles were evaluated by real time PCR. Libraries were sequenced on Novaseq 6000 using 1xSp-Flow (Illumina). A total of 130 to 210x106 paired reads were generated per sample.