Fig 1: NS1 binds at transcriptional regulatory elements of genes differentially expressed during RSV infection(A) Western blots show co-IP of FLAG-NS1 with MED1, MED14, and MED25. WCL, whole-cell lysate; Cy, cytoplasmic fraction; SN, soluble nuclear fraction; Ch, chromatin fraction.(B) Western blots of subcellular fractionated A549 cells expressing HA-NS1. U, unfractionated; M, membrane; Cy, cytosol.(C) Overlap of NS1 and Mediator ChIP-seq peaks in A549 cells.(D) Bar chart shows number of NS1 ChIP-seq peaks in functional genomic regions.(E) Heatmap shows differentially expressed genes with 10 kb of NS1 peaks at 24, 48, and 96 hpi with RSV (RNA-seq).(F) NS1 ChIP-seq peaks coincide with an enhancer (A), promoter (C), and Mediator peaks (B–D). Tracks: ChIP-seq as reads per million (RPM). Red and blue bars: called peaks (HOMER); GeneHancer track shows overlapping promoter (pink) and enhancers (green) (UCSC browser).(G) Bar graph shows fold change + standard deviation of luciferase activity of reporters with NS1-bound regions in the IFIT locus (A–D as in (F) relative to minimal promoter reporter (minP) in A549 cells transfected with empty vector (EV) or NS1 and treated with LMW polyI:C for 24 h. Unpaired t test, *p < 0.05, **p < 0.005, ns, not significant. All are representative of ≥2 experiments.