Fig. 1

GCLiPP sequencing reveals RNA transcript protein occupancy. A GCLiPP method of global RBP profiling. T cell RNAs are crosslinked to RBPs and lysates are biotinylated on primary amines. mRNAs are enriched with oligo-dT beads, and RBP-protected sites are digested, captured, sequenced, and aligned to the genome. B Film image of RBP-bound RNAs captured from Jurkats that underwent either UV crosslinking (UV 254 nm), protein biotinylation, or both. Lane marked “M” contains 19 and 24 nucleotide (nt) ssRNA ligated to radiolabeled 3′linker. RNA greater than 24nt + 3′linker size were extracted and processed for sequencing. C Normalized GCLiPP read depth (fraction of reads in called peak relative to all GCLiPP reads in annotated 3′ UTR) in two replicates of Jurkat cells. ρ represents Pearson correlation. D Proportion of mapped GCLiPP reads derived from genomic features. E Relative coverage of genomic features in GCLiPP sequencing reads relative to total length of genomic features of indicated class. F GCLiPP track of NR4A1 3′UTR. Red bars indicate presence of ARE motif (AUUUA). G GCLiPP track of IER3 gene along with predicted ROQUIN binding loop in the 3′UTR