Figure 7.

Comparison of the secondary structure of 7SL RNA within the Alu domain of SRP and SHAPE-guided structures of WT and mutated LTR78/AluJ RNAs. (a) Secondary structure of 7SL RNA within the Alu domain of SRP [65,66,99]. Helices (H) and loops (L) in orange are numbered according to a previously published topology [99]. The central stem (H3) is also known as helix 5 in the unifying nomenclature of all SRPs (circled) [67], stem V [68] or stem III [65]. Lower case letters indicate RNA sequences that are absent in the left Alu arm consensus [65] (Figure 5(d)). The C > U mutation is denoted by a black arrow. Blue and pink rectangles denote the central stem portions that are most similar to the F8 AluJ structure and were derived from the 5ʹ and 3ʹ parts of 7SL RNA, respectively. Conserved adenines with differential DMS reactivities in WT and mutated F8 probes are in red. (b,c) SHAPE-guided secondary structure of the WT (b) and mutant (c) LTR78/AluJ RNAs. Normalized DMS and NAI reactivities (colour coded as indicated) are shown in Figs. 5 and 6. Black triangles indicate major RNase A cleavage sites (numbered 1–7); their size indicates normalized cleavage intensities in the WT (Fig. S1). In the mutant (c), triangle sizes indicate changes in relative cleavage intensities as compared to the WT; red triangles denote >3-fold differences between the mutant and WT. The linkers and RT primers are highlighted in grey. Alternative structures are shown in Fig. S2. Decoy and active 5’ss are marked by closed and open green triangles, respectively. Secondary structures in panels (b) and (c) were predicted by RNAstructure [103] using normalized NAI constraints and default options. d Alignment of existing sense AluJ exons that employ the same decoy 5’ss as the F8 AluJ copy. The 5’ss are denoted by a slash. Dashes are alignment gaps; three orange dots separate the 5ʹ and 3ʹ parts of 7SL RNA that gave rise to Alu-like elements [30,66]. Full sequences of sense Alus that use 5’ss homologous to that activated in the AluJ copy in F8 are in Table S2. Their alignment is in Figure S3