Fig. 5: ADPRylation on its DBD restricts STAT1α binding to consensus motifs.

a Structure of DNA-bound STAT1α showing sites of ADPRylation on the DBD. STAT1α is shown in pink and DNA is shown in blue. ADPRylated residues (E393 and E394) are highlighted in red in the expanded view. The structure is from Protein Data Bank (PDB) 1BF5. b Heatmap of ChIP-seq data showing STAT1α enrichment in the top 50% of ‘gained’ STAT1α peaks in iBMDMs expressing wild-type (Wt) or DBD mutant (DBDmut) STAT1α with concurrent shRNA-mediated knockdown of endogenous STAT1α. The cells were treated ± IFNγ for 1 h. ‘Gained’ peaks were defined using a 4x MAD cutoff. c, d Browser tracks (c) and box plots (d) of ChIP-seq data representing ‘gained’ STAT1α peaks in iBMDMs expressing DBDmut compared to iBMDMs expressing Wt STAT1α (n = 227 peaks; Wilcoxon Signed-Rank test; p < 2.2 × 10^-16). Boxes represent 25^th–75^th percentile (line at median) with whiskers at 1.5*IQR. e Motifs enriched at gained STAT1α binding sites (DBDmut relative to Wt STAT1α). De novo motif analysis was performed using MEME. The predicted motifs were matched to known motifs using TOMTOM. P-values were generated using default parameters in TOMTOM (see Methods). f Binding of STAT1α DBDmut to non-consensus motifs. STAT1α from iBMDMs expressing Wt or DBDmut was incubated with double-stranded DNA oligonucleotides containing a consensus STAT3 binding sequence. Bound material and input were analyzed by immunoblotting for STAT1α. Uncropped immunoblots are provided as a Source Data file. g Gene expression associated with gained STAT1α peaks. Line plots representing fold change in nearest neighbor gene expression upon IFNγ treatment from RNA-seq in iBMDMs expressing Wt or DBDmut STAT1α with concurrent shRNA-mediated knockdown of endogenous STAT1α. h Line plots representing fold change in IFNγ-stimulated gene expression in iBMDMs expressing Wt or DBDmut STAT1α relative to an untreated control.