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. 2017 Nov 20;7:15833. doi: 10.1038/s41598-017-16177-9

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Inhibition of UPF1 or SMG-1 is required for generation of bulk of mutant protein from NMD-rescued PTC-containing gBglo-P66 mRNA. (a) Western analysis of siRNA mediated downregulation of core NMD and EJC factors UPF1, Y14, EIF4A3, SMG1, UPF2, and MAGOH in HeLa cells. GAPDH served as internal control. (b) qPCR analysis of relative β-globin mRNA levels in HeLa cells transfected with gBglo-WT or gBglo-P66 and siRNA against each factor. (c) Western analysis of β-globin derived from NMD-rescued PTC-containing gBglo-P66 mRNA in the same samples used in (b). (d) Concomitant inhibition of UPF1 with UPF2, Y14, EIF4A3, or MAGOH in HeLa cells transfected with gBglo-P66 in the presence and absence of MG132. Every experiment in Fig. 6 was performed two independent times and the results of one representative experiment are shown. Error bars in (a) represent the SD of the mean of two independently performed qPCR results.