Article

The efficiency of nonsense-mediated mRNA decay is an inherent character and varies among different cells

Department of Genetics, Life Sciences Institute, The Hebrew University, Jerusalem, Israel.
European Journal of HumanGenetics (Impact Factor: 4.23). 12/2007; 15(11):1156-62. DOI: 10.1038/sj.ejhg.5201889
Source: PubMed

ABSTRACT Nonsense-mediated mRNA decay (NMD) is a mechanism, which selectively degrades transcripts carrying premature termination codons (PTCs) and a variety of physiologic transcripts containing NMD-inducing features. In a recent study, we have found variable NMD efficiency among nasal epithelial cells obtained from cystic fibrosis (CF) patients. This variability was found for CF transmembrane conductance regulator (CFTR) transcripts carrying the W1282X PTC, as well as for several NMD physiologic substrates. Here, we aimed to investigate the possibility that variability in NMD efficiency is a more generalized phenomenon and is not restricted to nasal epithelial cells. To investigate this possibility, we analyzed the NMD efficiency of both a CFTR constructs carrying the W1282X PTC and beta-globin constructs carrying the NS39 PTC, in HeLa and MCF7 cells. Variability in NMD efficiency was found for both constructs between the cells, such that in HeLa cells the NMD was highly efficient and in MCF7 the efficiency was significantly lower. Moreover, similar differences in the efficiency of NMD were found for five endogenous NMD physiologic transcripts. Altogether, our results demonstrate existence of cells in which NMD of all transcripts is efficient, whereas others in which the NMD is less efficient, suggesting that the efficiency of NMD is an inherent character of cells. Our results also suggest that variability in the efficiency of NMD is a general phenomenon and is not restricted to nasal epithelial cells. As NMD affects the level of many transcripts, variability in the NMD efficiency might play a role as a genetic modifier of different cellular functions.

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    • "Beside the common F508del variant, our CF patients carry missense and nonsense alterations in the CFTR gene. Indeed, different studies have shown that the effect of nonsense mutations on CFTR mRNA levels are very variable [16] and nonsense-mediated decay (NMD) is addressed as the major mechanism for rapid degradation of aberrant transcripts harboring premature termination codons. Interestingly, microRNAs have recently been defined as important regulators of the NMD efficiency [17]. "
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    • "Gehring et al. reported that NMD can be triggered in a UPF2-inde- pendent manner [36], while Chan et al. describe an UPF3b-indepen- dent NMD pathway [37]. These studies highlight variants of the NMD mechanism and may explain why NMD efficiency varies significantly across different cell types [38]. Given the expression profile of ROD1 across different tissues, it is thus plausible that it could be another factor adding to the heterogeneity of NMD. "
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    • "If so, reducing the protein expression level should reduce or eliminate the complexes. HeLa cells are frequently used for studying nonsense mediated mRNA decay due to their low transfection efficiency and low levels of protein expression (Kang et al., 2009; Linde et al., 2007). Therefore we first determined the relative expression levels of wild-type γ2S subunits when coexpressed with α1 and β2 subunits in HeLa cells with the same amounts of cDNA used for HEK 293T cell transfections and measured subunit expression levels using the quantitative method of flow cytometry to compare subunit expression levels in each cell type (Figure 2A). "
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