Fig. 1: Folding of P1.1 helix upon Mn²⁺ binding stabilizes transcriptional pausing.

a The Mn²⁺-sensing riboswitch from L. lactis regulates gene expression at the transcriptional level. Stabilization of the P1.1 helix upon Mn²⁺ binding (Mn) prevents the formation of an intrinsic terminator hairpin, leading of the transcription of the downstream gene. Position of the G104 pause site is indicated as an orange star on the secondary RNA structure. b Representative denaturing gel showing the RNAP pauses during the transcription of the Mn²⁺ riboswitch. Position of the pause (G104), termination (T) and full-length (FL) products are indicated on the right. Experiments were performed using 25 µM rNTPs in the absence (−Mn²⁺) and presence (+Mn²⁺) of 0.5 mM Mn²⁺. The chase lanes (Ch) were taken at the end of the time course after an additional incubation with 500 µM rNTPs for 5 min. Unprocessed images are provided in the Supplementary Information. c Fraction of complexes at the G104 pause as a function of the transcription time in the absence (blue) and presence (orange) of 0.5 mM Mn²⁺. The reported errors are the SD of the mean from n = 2 independent replicates. d Quantification of the G104 pause half-life in the WT, MP1.1 and MP1.1cp riboswitch variants. Experiments were performed using 25 µM rNTPs in the absence (blue) and presence (orange) of 0.5 mM Mn²⁺. Error bars are the SD of the mean from n = 2 independent replicates. The statistical significance of differences was determined using the two-tailed Student’s t-test (***p < 0.01, **p < 0.05, *p < 0.1). The predicted conformation of the P1.1 is represented below each riboswitch variant dataset. Source data are provided with this paper at https://doi.org/10.7302/22513.