PEMT Couples Methionine Metabolism to Antiviral Defense by Restricting Influenza Virus NS1 mRNA Translation

A549 cells were infected by IAV (PR8, H1N1) for 12 hrs at MOI=1, and then collected A549 and crosslinked with 1% formaldehyde (10 min) and quenched with 125 mM glycine. Sonicated lysates (Bioruptor Pico, 15 cycles) were incubated overnight with 5 μg anti-PEMT, anti-eIF4G1, or isotype IgG antibodies pre-conjugated to Protein A/G magnetic beads. Following high-stringency washes and reverse crosslinking (65°C, 4 h), RNA was purified via phenol-chloroform extraction and quantified (Qubit). rRNA-depleted libraries (NEBNext Ultra II) were sequenced on an Illumina NovaSeq 4000 (150-bp paired-end). Raw reads were aligned to the human (hg38) and IAV (PR8) genomes using STAR; enriched regions were identified via MACS2 peak calling with an FDR < 0.05.