DNA transposons and alternative pre-mRNA splicing

DNA 转座子和选择性前 mRNA 剪接

• 批准号: 9926901
• 负责人: DONALD C RIO
• 金额: $ 65.26万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-01 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9926901
• 关键词: 5' Splice Site; Affect; Alternative Splicing; Animals; Behavior; Binding; Binding Sites; Cells; Complex; Courtship; DNA; DNA Binding Domain; DNA Transposable Elements; DNA Transposons; Defect; Disease; Drosophila genus; Elements; Exhibits; Family; Gene Expression; Gene Mutation; Genes; Genome; Genomics; Guanosine Triphosphate; Health; Human; Human Genome; Introns; Mobile Genetic Elements; Mus; N-terminal; Organism; Pattern; Phosphorus; Play; Polyadenylation; Primates; Process; Protein Isoforms; Protein Structure Initiative; Proteins; Proteomics; RNA; RNA Splicing; RNA-Binding Proteins; Rodent; Role; Split Genes; System; Tissues; Transcriptional Silencer Elements; Transposase; U1 Small Nuclear Ribonucleoprotein; Work; Xenopus; Zebrafish; base; cell type; cofactor; human embryonic stem cell; insight; mRNA Precursor; male; member; mutant; premature; public health relevance; transcriptome

 DESCRIPTION (provided by applicant): Mobile genetic elements or transposons are found in the genomes of all organisms. These elements can move via DNA or RNA intermediates. About 50% of the human genome is made up of transposable elements with ~ 2.7% corresponding to DNA-based transposons. Many of these putative transposons or transposase-related genes are uncharacterized. Our previous studies have focused on the P element family of DNA transposons in Drosophila. P element transposase functions as a tetramer, using GTP as a cofactor for transposition. N-terminal domain of the transposase corresponds to a C2CH THAP DNA binding domain, which is a member of a prevalent family of DNA binding domains found exclusively in animal genomes. One THAP gene, called THAP9, is homologous to the Drosophila P element transposase and is present in primates, Xenopus, zebrafish and Ciona, but is absent from rodents. Recent work from our lab has shown that the human and zebrafish THAP9 genes can mobilize the Drosophila and zebrafish P element transposons in human and Drosophila cells. This proposal is focused on understanding what role the human THAP9 gene may play in human embryonic stem cells and how the Drosophila P element transposase protein recognizes and assembles with the transposon ends, donor DNA, target DNA and GTP/Mg2+ to form an active protein-DNA complex. These studies are aimed at gaining mechanistic insights. Alternative pre-mRNA splicing is an important mechanism for regulating gene expression in metazoans and is a conduit through which genomic sequence is transferred to proteomic information. Most eukaryotic genes are split and have the potential for alternative splicing, dramatically increasing proteomic diversity. Many human and mouse disease gene mutations affect the splicing process. Splicing silencers are a major type of RNA control element generating tissue- or cell type-specific alternative splicing patterns. Our previous work has focused on characterization of the tissue-specific Drosophila P element pre-mRNA exonic splicing silencer element. Recent work from our group has focused on how the action of the RNA binding proteins, PSI and hrp48. Using this information, we want to identify new Drosophila cellular splicing silencer elements that are controlled by these two splicing factors. The PSI protein also interacts with U1 snRNP and PSI mutant Drosophila strains that abolish this interaction exhibit male courtship behavior defects and altered pre-mRNA splicing of the Drosophila male-specific fruitless pre-mRNA isoforms. We want to investigate how the PSI protein controls fruitless pre-mRNA splicing and how it controls binding of U1 snRNP on the Drosophila transcriptome. U1 snRNP has distinct roles in U1 snRNP binding sites in PCPA (premature cleavage and polyadenylation), splicing at intron 5' splice sites and at potential new splicing silencers.

 描述（由申请人提供）：在所有生物体的基因组中都发现了可移动遗传元件或转座子，这些元件可以通过 DNA 或 RNA 中间体移动。大约 50% 的人类基因组由转座元件组成，其中约 2.7% 对应于转座子。基于 DNA 的转座子。我们之前的研究主要集中在果蝇 P 元件的 P 元件家族。转座酶以四聚体的形式发挥作用，使用 GTP 作为转座酶的 N 端结构域，对应于 C2CH THAP DNA 结合结构域，它是仅在动物基因组中发现的常见 DNA 结合结构域家族的成员。 THAP9 基因与果蝇 P 元件转座酶同源，存在于灵长类动物、爪蟾、斑马鱼和海鞘中，但在啮齿类动物中不存在。我们实验室的研究表明，人类和斑马鱼的 THAP9 基因可以在人类和果蝇细胞中动员果蝇和斑马鱼的 P 元件转座子。该提案的重点是了解人类 THAP9 基因在人类胚胎干细胞中可能发挥的作用以及果蝇如何发挥作用。 P元件转座酶蛋白识别并与转座子末端、供体DNA、靶DNA和GTP/Mg2+组装，形成活性蛋白-DNA复合物。选择性前mRNA剪接是调节后生动物基因表达的重要机制，也是将基因组序列转移到蛋白质组信息的渠道，大多数真核基因被分裂并具有选择性剪接的潜力，从而显着增加。蛋白质组多样性。许多人类和小鼠疾病基因突变影响剪接过程，剪接沉默子是产生组织或细胞类型特异性选择性剪接模式的主要类型。我们小组最近的工作重点是组织特异性果蝇 P 元件前 mRNA 外显子剪接沉默元件的表征，我们希望利用这些信息来鉴定新的果蝇。由这两个剪接因子控制的细胞剪接沉默元件元件还与 U1 snRNP 和 PSI 突变果蝇菌株相互作用，消除了这种相互作用，表现出雄性求爱行为缺陷并改变。果蝇雄性特异性无结果前 mRNA 亚型的前 mRNA 剪接 我们想要研究 PSI 蛋白如何控制无结果前 mRNA 剪接，以及它如何控制 U1 snRNP 在果蝇转录组上的结合，在 U1 中具有独特的作用。 PCPA 中的 snRNP 结合位点（过早切割和聚腺苷酸化）、内含子 5' 剪接位点处的剪接以及潜在的新剪接位点拼接消音器。