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. Author manuscript; available in PMC: 2011 May 3.
Published in final edited form as: Cell Res. 2008 Sep;18(9):921–936. doi: 10.1038/cr.2008.66

Figure 5. AU-constitution of the upstream stem-loop I is important for producing proteins initiated from the GGC1274–1276 codon.

Figure 5

(A) Schematic representation of the partial ACAT1 mRNA sequence (nt 1243–1786) and mutations of stem-loop I. The detailed sequence of nt 1253–1270 is listed without or with the mutant nucleotides (underlined). Gray bar, partial ACAT1 mRNA sequence (ACAT1 1243–1786); black bar, 3×Flag coding sequence (3×Flag); filled circle, GGC1274–1276 initiation codon; hollow circle, AUG1397–1399 initiation codon.

(B) The expression plasmids depicted in (A) are transiently transfected into AC29, the lysates are prepared and immunoblotting is carried out with anti-ACAT1 antibodies (DM10). Arrows indicate the positions of ACAT1-NT-Flag proteins respectively initiated from GGC1274–1276 and AUG1397–1399. The experiments are repeated twice with similar results.

(C) Predicted RNA secondary structures of nt 1253–1270 without or with mutations of stem-loop I depicted in (A). The folding Gibbs free energy (ΔG) of predicted RNA secondary structure is listed beneath each one. These experiments are repeated three times with similar results.