Functional analysis and characterization of protein-binding sites. (A) Distribution of ssRNase-treated (light blue bars) and dsRNase-treated (green bars) PPS sizes from formaldehyde-cross-linked samples. Dashed lines represent median PPS sizes (ssRNase, blue line and dsRNase, green line). (B) Genomic distribution of PPS density, measured as PPS base coverage normalized to RNase digestion control read counts per genomic region. Proximal intron refers to 500 nucleotides at the 5′ and 3′ ends of introns. (C) Cumulative distribution of average SiPhy-π scores in PPSs (red line) versus similarly sized flanking sequences (gray line). (D) Comparison of average SiPhy-π scores between PPSs (red bars) and flanking sequences (gray bars) for various genomic regions. (E) Average SiPhy-π score profiles across the first and last 25 nucleotides of PPSs as well as 50 nucleotides upstream and downstream of exonic (green line), intronic (blue line) and lncRNA (orange line) PPSs. *** denotes P < 2.2 × 10–16 (chi-squared test). CDS, coding sequence; dsRNase, double-stranded RNase; lncRNA, long non-coding RNA; NS, not significant; PPS, protein-protected site; ssRNase, single-stranded RNase; UTR, untranslated region.