Isolation of proteins binding to precursor miRNA terminal loops. (A) Predicted structures of apical regions with highlighted putative protein-binding sites for let-7a-1 and miR-101-1. (B) Standard and (C) RNase-assisted RNA chromatography of let-7a-1 and miR-101-1 terminal loops in HeLa nuclear extracts. Bands denoted A and B correspond to selected proteins highly enriched in the let-7a-1 chromatograms, hnRNP L, and nucleolin, respectively. Bands C and D correspond to PTB protein, which is highly enriched in the miR-101-1 chromatograms. RNase-assisted RNA chromatography was performed using RNase A/T1 or RNase I, as indicated. (D) RNase-assisted RNA chromatography with let-7a-1 TL and increasing concentration of RNase A/T1 (1, 2.5, and 5 μL per reaction).
