RIP Assay for YTHDF2 Binding to EBV Genes

RIP was conducted using the Magna RIP kit (Cat #17–700, Millipore) following the manufacturer’s instructions. In brief, Akata (EBV+) cells expressing YTHDF2-Myc or YTHDF2 (K281R/K571R/K572R, RRR)-Myc were treated with IgG cross-linking for 24 h and then lysed using the RIP lysis buffer provided in the kit. A portion of the lysate (10%) was saved as the input sample. The anti-c-Myc magnetic beads (Cat #88842, Thermo Scientific) were washed with RIP buffer and incubated with RNA overnight at 4°C. On the following day, the beads were collected and washed six times with the RIP wash buffer. The enriched RNA-protein complex was digested with proteinase K, and the released RNA was purified using phenol-chloroform extraction. The purified RNA was then subjected to reverse transcription for subsequent qPCR analysis using the primers for ZTA, RTA BALF5, BGLF4, BLLF1, and MALAT1 as we previously described (7 (link), 13 (link))