Fig. 1

Nuc-SHAPE-Structure-Seq method can accurately probe the in vivo RNA structure of nuclear RNAs. a Schematic pipeline of Nuc-SHAPE-Structure-Seq for both in vivo and deproteinized conditions. Asterisks, SHAPE modification; blue oval, protein; RT, reverse transcription. For in vivo treatments (left), NAI was applied to Arabidopsis thaliana seedlings directly and single-stranded nucleotides of RNA were modified. SHAPE treatment was also applied on the RNAs after nuclei isolation and protein removal, which we termed the “deproteinized condition” (right). Deep sequencing was performed followed by the RT-stop counting. b SHAPE reactivity profiles of U1 and U12 snRNAs. SHAPE reactivity profiles of both in vivo (blue) and deproteinized (orange) conditions were shown. Double-stranded regions were shaded with gray. Sm protein binding sites were highlighted with yellow boxes. At Sm protein binding sites, significantly higher SHAPE reactivities were observed under the deproteinized condition rather than the in vivo condition for both U1 and U12 snRNAs (paired t test, P value = 6.8e−3 and 3.1e−6 for U1 and U12 snRNA, respectively). c SHAPE reactivities are consistent with the phylogenetically derived U1 and U12 snRNA structures. Sm protein binding sites were highlighted with black boxes. Nucleotides were color-coded according to in vivo and deproteinized SHAPE reactivity values (SHAPE reactivity 0.6–1.0 marked in red, 0.3–0.6 marked in yellow, 0–0.3 marked in green)