Human ribosomes from cells with reduced dyskerin levels are intrinsically altered in translation
Human ribosomes from cells with reduced dyskerin levels are intrinsically altered in translation
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DOI：
10.1096/fj.15-270991
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发表时间：
2015-08-01
期刊：
影响因子：
4.8
通讯作者：
Montanaro, Lorenzo
中科院分区：
文献类型：
--
作者：
Penzo, Marianna;Rocchi, Laura;Montanaro, Lorenzo
Dyskerin是一种假尿苷（psi）合酶，参与基本的细胞过程，包括rRNA和小核RNA中的尿苷修饰和端粒稳定。在X连锁先天性角化不良（X-DC）和癌症中，Dyskerin功能发生改变。Dyskerin在rRNA假尿苷酸化中的作用被认为是缺乏dyskerin功能的细胞中所描述的mRNA翻译改变的基础，尽管目前缺乏相关的直接证据。我们的目的是明确地确定是否有缺陷的dyskerin功能可能决定一个内在的核糖体缺陷，导致合成活性的改变。因此，将来自dyskerin耗尽的人细胞的核糖体纯化，并1）添加到不含核糖体的受控网织红细胞无细胞系统中以研究mRNA翻译; 2）通过质谱分析蛋白质污染和组成，3）分析整体假尿苷酸化水平。从不良角蛋白缺失细胞中纯化的核糖体显示出翻译保真度的改变和内部核糖体进入位点（IRES）介导的翻译。这些核糖体显示减少尿苷修饰，而他们没有不同的蛋白质污染或核糖体蛋白质组成方面的核糖体从匹配的对照细胞与完整dyskerin活性。总之，人类细胞中dyskerin功能的缺乏诱导rRNA尿苷修饰的缺陷，这足以改变核糖体活性。Penzo，M.，罗基湖，Brugiere，S.，Carnicelli，D.，Onofrillo角，Coute，Y.，Brigotti，M.，蒙塔纳罗湖来自dyskerin水平降低的细胞的人核糖体在翻译中固有地改变。
Dyskerin is a pseudouridine (psi) synthase involved in fundamental cellular processes including uridine modification in rRNA and small nuclear RNA and telomere stabilization. Dyskerin functions are altered in X-linked dyskeratosis congenita (X-DC) and cancer. Dyskerin's role in rRNA pseudouridylation has been suggested to underlie the alterations in mRNA translation described in cells lacking dyskerin function, although relevant direct evidences are currently lacking. Our purpose was to establish definitely whether defective dyskerin function might determine an intrinsic ribosomal defect leading to an altered synthetic activity. Therefore, ribosomes from dyskerin-depleted human cells were purified and 1) added to a controlled reticulocyte cell-free system devoid of ribosomes to study mRNA translation; 2) analyzed for protein contamination and composition by mass spectrometry, 3) analyzed for global pseudouridylation levels. Ribosomes purified from dyskerin-depleted cells showed altered translational fidelity and internal ribosome entry site (IRES)-mediated translation. These ribosomes displayed reduced uridine modification, whereas they were not different in terms of protein contamination or ribosomal protein composition with respect to ribosomes from matched control cells with full dyskerin activity. In conclusion, lack of dyskerin function in human cells induces a defect in rRNA uridine modification, which is sufficient to alter ribosome activity.-Penzo, M., Rocchi, L., Brugiere, S., Carnicelli, D., Onofrillo, C., Coute, Y., Brigotti, M., Montanaro, L. Human ribosomes from cells with reduced dyskerin levels are intrinsically altered in translation.