Regulation and Function of snoRNA Genes

snoRNA基因的调控和功能

• 批准号: 8755989
• 负责人: Wendy Victoria Gilbert
• 金额: $ 26.74万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8755989
• 关键词: Affect; Amino Acids; Biological Process; Biology; Breast; Cancer Patient; Cells; Cellular Stress; Codon Nucleotides; Colorectal Cancer; Data; Enzymes; Eukaryota; Functional RNA; Functional disorder; Future; Genes; Genetic; Genetic Code; Guide RNA; Health; Heat-Shock Response; Human; Lead; Length; Malignant Neoplasms; Malignant neoplasm of brain; Malignant neoplasm of lung; Malignant neoplasm of prostate; Maps; Mass Spectrum Analysis; Messenger RNA; Methods; Methylation; Modeling; Modification; Molecular Target; Multiple Myeloma; Nuclear; Nucleotides; Nutrient; Organism; Oxidative Stress; Pattern; Positioning Attribute; Pseudouridine; Reading; Regulation; Ribosomal RNA; Ribosomes; Risk; Sense Codon; Site; Small Nucleolar RNA; Specificity; Techniques; Terminator Codon; Testing; Therapeutic; Time; Translating; Translations; Treatment-Related Cancer; United States; Validation; Woman; Work; Yeasts; deprivation; design; environmental change; genome-wide; killings; leukemia; men; mutant; public health relevance; research study; response

DESCRIPTION (provided by applicant): snoRNAs are highly conserved non-coding RNAs that direct two types of site-specific post-transcriptional modifications of other RNAs - pseudouridinylation and 2'-O-methylation. Dysregulation of specific snoRNAs is associated with increased risk for breast, prostate, lung, brain and colorectal cancers as well as leukemia and multiple myeloma, but the mechanistic connections between altered snoRNA function and cancer are not understood. Recent discoveries herald a paradigm shift in the snoRNA field. First, snoRNAs were shown to have a greatly expanded list of potential RNA targets, including mRNAs. Second, some snoRNA-directed modifications were shown to be regulated by environmental changes. Finally, the presence of pseudouridine was shown to change how the genetic code is interpreted by ribosomes. Together, these results suggest potentially widespread regulatory functions for snoRNAs through directing post-transcriptional modification of mRNAs. We hypothesize that snoRNAs direct physiologically relevant pseudouridinylation of mRNA targets by a mechanism that involves relaxed specificity pairing between guide sequence and target RNA. To test this hypothesis, we have developed an efficient method for mapping the sites of snoRNA-directed pseudouridinylation genome-wide with single nucleotide precision (Pseudo-seq). Our preliminary results from Pseudo-seq experiments provide the first evidence that endogenous mRNAs are pseudouridinylated. Thus, we are uniquely well positioned to investigate this new paradigm for snoRNA function. In Aim 1 we will identify the regulated targets of snoRNA-directed pseudouridinylation in response to nutrient deprivation, oxidative stress and heat shock using Pseudo-seq. Mutants in the catalytic component of snoRNA enzymes, Cbf5/Dyskerin, will be used to determine which pseudouridines are formed by a snoRNA-dependent mechanism. Completion of this aim will comprehensively identify mRNA targets of regulated snoRNA-dependent modification in yeast. Aim 2 will use computational and targeted genetic approaches to dissect the sequence requirements for regulation by snoRNAs. Completion of this aim will facilitate accurate prediction of snoRNA targets in all eukaryotic organisms. Finally, Aim 3 will use mass spectroscopy to determine the functional consequences of mRNA pseudouridinylation for decoding by the ribosome. Completion of this aim has the potential to reveal substantial rewiring of the genetic code. Human health relevance: The results of this study will reveal new biological functions of snoRNAs and thereby illuminate the potential mechanisms by which snoRNA dysfunction leads to cancer. We believe that in the future, a better understanding of the function of snoRNAs in cells may lead to better treatment of cancers associated with perturbation of the function of snoRNA genes.

描述（由申请人提供）：SNORNAS是高度保守的非编码RNA，指导两种类型的位点特异性转录后修饰 - 其他RNA-假胞胺丁基化和2'-O-甲基化。特定snornas的失调与增加乳腺，前列腺，肺，大脑和结直肠癌以及白血病和多发性骨髓瘤的风险增加有关，但是尚不清楚SINORNA功能和癌症改变的机械连接。最近的发现预示着Snorna领域的范式转变。首先，SNORNA的潜在RNA靶标有很大扩展的列表，包括mRNA。其次，一些snorna导向的修改被证明受环境变化的调节。最后，证明假氨酸的存在会改变核糖体解释遗传代码的方式。总之，这些结果表明，通过指导mRNA的转录后修饰，可能会广泛地调节性能。我们假设SNORNA通过一种涉及指导序列和靶RNA之间宽松的特异性配对的机制直接对mRNA靶标的生理相关的假素化。为了检验这一假设，我们开发了一种有效的方法来绘制以单核苷酸精度（伪seq）的snoRNA定向伪苷基因组的位点。我们的伪seq实验的初步结果提供了第一个证据，表明内源性mRNA是伪丁素化的。因此，我们在研究SNORNA功能的这一新范式方面处于独特状态。在AIM 1中，我们将通过使用伪seq来确定响应营养剥夺，氧化应激和热休克的副类固定素化的调节靶标。 SNORNA酶（CBF5/DYSKERIN）催化成分中的突变体将用于确定哪些假源是由SNORNA依赖性机制形成的。该目标的完成将全面识别酵母中依赖SNORNA依赖性修饰的mRNA靶标。 AIM 2将使用计算和靶向遗传方法来剖析SNORNA的调节序列要求。该目标的完成将有助于准确预测所有真核生物中的SNORNA靶标。最后，AIM 3将使用质谱法来确定mRNA伪啶基化对核糖体解码的功能后果。该目标的完成有可能揭示遗传密码的重大重新布线。人类健康相关性：这项研究的结果将揭示SNORNAS的新生物学功能，从而阐明SNORNA功能障碍导致癌症的潜在机制。我们认为，将来，对snoRAS在细胞中的功能的更好理解可能会导致更好地治疗与snorna基因功能相关的癌症。