Anczuków Olga, Ware Mark D, Buisson Monique, Zetoune Almoutassem B, Stoppa-Lyonnet Dominique, Sinilnikova Olga M, Mazoyer Sylvie
Laboratoire de Génétique Moléculaire, Signalisation et Cancer UMR5201 CNRS, Université Lyon 1, Lyon, France.
Hum Mutat. 2008 Jan;29(1):65-73. doi: 10.1002/humu.20590.
The nonsense-mediated mRNA decay (NMD) mechanism is an evolutionarily conserved process ensuring the degradation of transcripts carrying premature termination codon(s). NMD is believed to prevent the synthesis of truncated proteins that could be detrimental to the cell. However, although numerous studies have assessed the efficiency of this mechanism at the mRNA level, data are lacking in regard to whether NMD fulfills its expected goal at the protein level. In this study, we have investigated whether endogenous alleles of breast cancer predisposing genes carrying nonsense codons were able to produce detectable amounts of truncated proteins in lymphoblastoid cell lines. A total of 20 truncating BRCA1 mutations were analyzed, along with the 1100delC CHEK2 and the 770delT TP53 mutations. All the studied alleles triggered NMD, the amount of mutant transcript ranging from 16 to 63% of that of the wild-type species. We found that BRCA1 and CHK2 truncated proteins could not be detected, even when NMD was inhibited. This suggests that BRCA1 and CHK2 truncated proteins are highly unstable. Conversely, the p53 protein encoded by the 770delT allele is as abundant as the wild-type protein, as removal of the C-terminal p53 domain leads to a stabilized mutant protein, whose abundance is markedly increased when NMD is inhibited. Therefore, our results show that it is not possible to infer the presence of truncated proteins in cells from carriers of a truncated mutation without experimental verification, as each case is expected to be different.
无义介导的mRNA衰变（NMD）机制是一个进化上保守的过程，可确保降解携带过早终止密码子的转录本。人们认为NMD可防止可能对细胞有害的截短蛋白的合成。然而，尽管众多研究已在mRNA水平评估了该机制的效率，但关于NMD在蛋白质水平是否实现其预期目标的数据仍然缺乏。在本研究中，我们调查了携带无义密码子的乳腺癌易感基因的内源性等位基因是否能够在淋巴母细胞系中产生可检测量的截短蛋白。总共分析了20个截短的BRCA1突变，以及1100delC CHEK2和770delT TP53突变。所有研究的等位基因均触发NMD，突变转录本的量为野生型物种的16%至63%。我们发现，即使抑制NMD，也检测不到BRCA1和CHK2截短蛋白。这表明BRCA1和CHK2截短蛋白高度不稳定。相反，由770delT等位基因编码的p53蛋白与野生型蛋白一样丰富，因为去除C末端p53结构域会导致突变蛋白稳定，当NMD被抑制时其丰度会显著增加。因此，我们的结果表明，在没有实验验证的情况下，不可能从截短突变携带者的细胞中推断出截短蛋白的存在，因为预计每种情况都不同。
