X-linked Dyskeratosis Congenita and ribosomal frameshifting

描述(申请人提供)：与人类先天性和获得性骨髓衰竭综合征有关的核糖体功能异常被归类为核糖体疾病的广泛保护伞。我们两个实验室最近的合作开启了一条全新的研究途径，通过建立伪尿苷合成酶Dyskerin介导的核糖体修饰在翻译控制的特定方面的进化保守作用。伪尿嘧啶修饰的功能作用具有重要的医学意义，因为编码dyskerin的基因DKC1的突变在许多疾病中都被发现，包括X-连锁先天性角化不良(X-DC)、Hoyeraal-Hreidarsson综合征和许多癌症。重要的是，我们发表的工作已经发现，rRNA缺陷核糖体对tRNAs的亲和力降低导致程序性-1核糖体框架移位率(-1PRF)增加，这是一种正在成为基因表达重要调节因素的分子机制。这一建议试图检验一种假设，即-1 PRF的全球变化影响编码关键造血因子的特定mRNAs亚集的表达，这可能有助于X-DC的一些病理特征。我们广泛的已发表和未发表的研究结果明显支持这一假说，这些结果表明，-1 PRF的增加通过无意义介导的mRNA衰退(NMD)途径促进特定mRNAs的快速降解。此外，我们还证明了携带不同DKC1突变的X-DC患者的造血细胞中rRNA水平受损。重要的是，我们的初步和发表的数据也表明，参与端粒维持的一类重要的mRNAs在酵母和人类细胞中都受到这一机制的调节，并在X-DC患者中被破坏，从而将rRNA缺陷和端粒缩短联系在一起。通过利用X-DC作为疾病范例，并结合三个模型系统，酵母、小鼠和人类，在这个建议中，我们将扩展我们的新发现，识别与核糖体功能障碍相关的骨髓衰竭背后的mRNAs谱系，并确定在X-DC中抑制NMD和-1 PRF的治疗潜力。这项建议的目标1试图使用两种互补的方法来确定其表达受到酵母、小鼠和人类细胞中rRNA缺陷影响的mRNAs谱系。目的2将确定受损的rRNA改变含有mRNAs的-1PRF表达的机制。目的3旨在确定靶向NMD和-1PRF在X-DC中的治疗潜力。到拟议的研究结束时，我们将1)建立一个理解核糖体疾病的新范式，2)确定可用作生物标记物并作为治疗干预目标的特定基因，3)探索治疗这类疾病的新方法。

DESCRIPTION (provided by applicant): Abnormalities in ribosome function that are implicated in both congenital and acquired bone marrow failure syndromes in humans are classified under the broad umbrella of ribosomopathies. A recent collaboration between our two laboratories has initiated a completely new avenue of research by establishing an evolutionarily conserved role for ribosome modifications mediated by the pseudouridine synthase dyskerin in specific aspects of translation control. The functional role of pseudouridine (¿) modifications is of great medical importance as mutations in DKC1, the gene encoding for dyskerin, are found in a number of diseases including X-linked dyskeratosis congenita (X-DC), Hoyeraal-Hreidarsson syndrome, and numerous cancers. Importantly, our published work has identified that the decreased affinity of rRNA ¿ defective ribosomes for tRNAs results in increased rates of programmed -1 ribosomal frameshifting (-1 PRF), a molecular mechanism that is emerging as an important regulator of gene expression. This proposal seeks to test the hypothesis that global changes in -1 PRF affect the expression of specific subsets of mRNAs encoding key hematopoietic factors, that may contributing to some of the pathological features associated with X- DC. Support for this hypothesis is evident from our extensive published and unpublished findings showing that increased rates of -1 PRF promotes rapid degradation of specific mRNAs through the Nonsense-Mediated mRNA Decay (NMD) pathway. Furthermore, we have demonstrated that rRNA ¿ levels are impaired in hematopoietic cells from X-DC patients harboring distinct DKC1 mutations. Importantly, our preliminary and published data also suggest that one important functional class of mRNAs involved in telomere maintenance is regulated by this mechanism in both yeast and human cells and is disrupted in X-DC patients, thus linking rRNA ¿ defects and telomere shortening. By utilizing X-DC as a disease paradigm and combining three model systems, yeast, mouse and humans, in this proposal we will extend our novel findings and identify the repertoire of mRNAs that underlie bone marrow failure associated with ribosome dysfunction and determine the therapeutic potential of inhibiting NMD and -1 PRF in X-DC. Aim 1 of this proposal seeks to identify the repertoire of mRNAs whose expression is affected by rRNA ¿ defects in yeast, mouse and human cells using two complementary approaches. Aim 2 will determine the mechanisms through which impaired rRNA ¿ alters expression of -1 PRF containing mRNAs. Aim 3 is oriented towards determining the therapeutic potential of targeting NMD and -1 PRF in X-DC. By the end of the proposed studies, we will have 1) established a new paradigm for understanding ribosomopathies, 2) identified specific genes that can be used as biomarkers and targeted for therapeutic intervention, and 3) explored novel approaches to treat this class of diseases.