Figure 4. Force Applied Along RNA Affects TE and an Energetic Model of Termination.

(A)-(C) Termination efficiency (mean ± SEM) plotted as a function of the load applied between the nascent RNA and RNAP. (A) The complete his terminator (dark blue) and associated U-tract alone (light blue), (B) the complete t500 terminator (dark green) and associated U-tract alone (light green), (C) the complete tR2 terminator (dark red) and associated U-tract alone (light red). For each complete terminator, two values for TE at F = 0 are plotted: the TE determined in bulk solution using a gel-based assay (black filled circles) and the TE measured in the same fashion, but in the presence of a complementary oligo that prevents secondary structure from forming in the upstream region (the endogenous TE, shown by the open, colored circle at F = 0). Solid light-colored lines show global fits to Equations 2 & 4, solid dark-colored lines show fits to Equations 2 & 5. (D) An energy landscape representation of the quantitative model for termination. The TE is determined by the difference, ΔE_total, between energy barriers to elongation (right) and termination (left). The portion of the landscape depicted in red represents the barrier to termination for terminator U-tracts, which can be modulated by force on the RNA, lowering it by ~F ×·δ_hybrid (dotted red line; the exact expression for modulation of this barrier includes corrections for the mechanical properties of ssRNA, not shown here; see Supplemental Materials and Equation 4) The inclusion of a hairpin stem lowers the termination barrier by E_stem (solid green line). Force on RNA biases against folding of the terminator hairpin, raising the barrier to termination by ~F ×·δ_stem (dotted green line). The overall TE is related to ΔE_total through Equation 2. (E) Table of global best-fit parameters, obtained by fitting the TE for all three terminators and associated U-tracts at all forces to Equation 2 & 5.