ATAC-seq Profiling of HTLV-1 Infected Cells

ATL patient PBMCs were thawed and washed with PBS containing 0.1% BSA. To discriminate dead cells, we used the LIVE/DEAD Fixable Dead Cell Stain Kit (Invitrogen). For cell surface staining, cells were stained with APC anti-human CD4 (clone: RPA-T4) (BioLegend) and anti-SynCAM (TSLC1/CADM1) mAb-FITC (MBL) antibodies for 30 minutes at 4°C followed by a wash with PBS. HTLV-1 infected cells (CADM1+ and CD4+) were sort-purified with FACS Canto (Beckman Coulter) to reach 98–99% purity. Data was analyzed by FlowJo software (Treestar). Soon after the sorting, 10000-50000 HTLV-1 infected cells were centrifuged and used for ATAC-seq as previously described [5 (link)]. Total RNA was isolated from the remaining cells using the RNeasy Mini Kit (Qiagen). Library preparation and high-throughput sequencing were performed by Macrogen Inc. (Seoul, Korea). The diagnostic criteria and classification of clinical subtypes of ATL were performed as previously described [28 ]. 77 ATAC-seq datasets from 13 human primary blood cell types and datasets from 42 AML patients were obtained from the Gene Expression Omnibus (GEO) with accession number GSE74912 [7 (link)]. ATAC-seq datasets from 7 CLL patients were obtained from GSE111015 [18 (link)] and RNA-seq datasets of CD4⁺T and Mono cells were obtained from GSE74246 [7 (link)].