Figure 5.

Cap-distal, but not cap-proximal, SL insertion confers strong Ded1 dependence on LUC reporter expression. (A) Schematics of LUC reporters containing promoter, 22-nt 5′ UTR, and first four codons of RPL41A fused to FLUC coding sequences and modified RPL41A 3′ UTR, plus indicated 5′ UTR insertions of CAA repeats. LUC expression was assayed as in Figure 4B and mean (±SEM) ratios of expression in ded1-cs versus WT cells are plotted. (B) Schematics to indicate the position and stability of SLs (predicted at http://rna.urmc.rochester.edu/RNAstructure.html) inserted into the 91-nt 5′ UTR reporter in A. LUC expression was assayed as in Figure 4B; and mean (±SEM) expression (in million units) in WT (middle) and ratios of expression in ded1-cs versus WT cells (right) are plotted. (C) Same as B except that LUC expression was measured in three transformants of WT and tif1-ts by preculturing in SC−Leu−Ura at 23°C, diluting to OD₆₀₀∼0.1, and growing for ∼three doublings at 33°C (∼24 h for tif1-ts and ∼6 h for WT).