Figure 5. in vitro vs. in-cell SHAPE-Seq reactivity map comparisons for 5S rRNA and the TPP riboswitch.

(A) 5S rRNA in-cell reactivities overlaid on a predicted secondary structure [72] and a three dimensional model of the 5S rRNA within the entire ribosome (inset; from PDB 4V69) [73]. Individual ribosomal proteins (L5, L18, L25, L27) and the 23S rRNA are labeled on the secondary structure near their approximate locations; helices are numbered I–V. Reproduced from Watters et al., 2015 [26] with permission from Oxford University Press. (B) Comparison of reactivities for the E. coli 5S rRNA measured in-cell (endogenous expression, top, RMDB: 5SRRNA_1M7_0007) vs. in vitro (bottom, RMDB: 5SRRNA_1M7_0009). Reactivities are color-coded according to (A). Clear differences in the endogenous 5S rRNA reactivities are apparent, especially for nucleotides 35–38 and 44–100, which increase and decrease, respectively relative to in vitro. (C) Comparison of the E. coli thiM TPP riboswitch measured in vivo (expressed from a plasmid vector, top, RMDB: TPPSC_1M7_0004) vs. an in vitr measurement of the adapter domain only with 5 μM TPP present (RMDB: TPPSC_1M7_0005). Comparing the reactivities in the 5′ half of the aptamer domain suggests that the riboswitch is primarily in the bound form in the cell, though differences in the 3′ half suggest that the cellular environment and the aptamer sequence context affect the RNA fold. Nucleotides that were not mapped in (B) and (C) are indicated with gray.