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. 2011 Jun 25;39(17):7598–7609. doi: 10.1093/nar/gkr484

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© The Author(s) 2011. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 6. — A. The influence of eIF1 on translation initiation in vitro from short 5′ UTR mRNA and distinct-AUG contexts. (A) schematic representation of the GFP-reporter gene with either TISU or a strong AUG context, both with 11 nucleotides, 5′ UTR. eIF1 was expressed in E. coli, purified and the indicated amounts were added to in vitro-translation reactions with the described constructs. Reactions with TISU and the strong AUG context are indicated at the top. (B) The effect of eIF1 on translation efficiency and accuracy in vivo. The upper panel shows a scheme of the GFP-reporter gene driven either by TISU or by a weak-AUG context, both with short 5′ UTR. The AUG-flanking sequence is shown. These reporters were transfected into HEK293T cells with increasing amounts of eIF1-expression plasmid as indicated, and the translation-initiation site was determined by immunoblot with GFP-specific antibody. US and DS denote upstream and downstream initiation site, respectively. eIF1 expression was analyzed by immunoblot using anti-HA antibody. (C) A graph representing translation directed by TISU or the weak-AUG reporter, GFP, in the absence or presence of low dose of eIF1 plasmid (250 ng), from three independent experiments (average ± SD). The overall translation without eIF1 was set to one. The relative intensity of the upstream translation site is presented by light grey bars and the DS-translation site by dark grey bars. The asterisks denotes statistically significant difference, P < 0.005.