Figure 6.

Changing levels of MiaA increase frameshifting. (A) Diagram depicts the structures of the dual luciferase reporters, with specific intergenic linker sequences and premature stop codons (red) indicated below. Each linker contains multiple MiaA-sensitive UNN codons (underlined). (B and C) Graphs show results from + 1 and –1 frameshifting assays with UTI89, UTI89ΔmiaA, UTI89/pRR48, or UTI89/pMiaA_Ptac carrying one of the dual luciferase reporter constructs. Bacteria were grown shaking at 37°C in LB, with 1 mM IPTG included for UTI89/pRR48 and UTI89/pMiaA_Ptac. After reaching an OD₆₀₀ of ∼0.2, 0.2% arabinose was added to induce expression of the luciferases. At an OD₆₀₀ of 0.5, translational error rates were quantified by determining the ratio of firefly to renilla luciferase activities in bacteria carrying the +1 (Az1) and –1 (HIV) reporter constructs. Results were normalized using the ratio of firefly to renilla luciferase activity in bacteria carrying control plasmids in which the luciferases are in-frame. **P< 0.01; ****P < 0.0001 by two-tailed unpaired t tests; n = 10–14 independent replicates.