Repeat-containing RNA Binding Proteins of Trypanosomes

含有重复序列的锥虫 RNA 结合蛋白

• 批准号: 8884913
• 负责人: Inna Afasizheva
• 金额: $ 40.93万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-01-05 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8884913
• 关键词: Action Potentials; Address; African Trypanosomiasis; Amino Acids; Animal Model; Appearance; Area; Binding; Biochemical; Biochemistry; Biogenesis; Biological Assay; Biology; Code; Complex; Country; Crystallography; DNA Structure; Deposition; Disadvantaged; Disease; Drug Targeting; Drug resistance; Due Process; Ensure; Enzymes; Event; Family; Gene Expression; Gene Expression Profile; Genetic; Genome; Goals; Guide RNA; Health Hazards; Helix-Turn-Helix Motifs; Human; Investigation; Left; Mass Spectrum Analysis; Mastigophora; Mediating; Messenger RNA; Mitochondria; Mitochondrial RNA; Modification; Molecular Biology; Monitor; Nature; Nuclear; Open Reading Frames; Organism; Parasites; Parasitology; Pathway interactions; Phosphodiesterase I; Polyadenylation; Polynucleotide Adenylyltransferase; Process; Protein Engineering; Proteins; Proteomics; Publishing; RNA; RNA Binding; RNA Editing; RNA Processing; RNA Sequences; RNA-Binding Proteins; Reaction; Recruitment Activity; Repression; Research; Ribosomal RNA; Ribosomes; Role; Shapes; Small RNA; Source; Specificity; Structure; Tail; Testing; Therapeutic; Therapeutic Intervention; Transcript; Translations; Trypanosoma; Trypanosoma brucei brucei; Work; base; blocking factor; crosslink; deep sequencing; health economics; in vivo; insertion/deletion mutation; mRNA Decay; mRNA Precursor; mRNA Stability; member; public health relevance; resistant strain; transcriptome sequencing; vector control

 DESCRIPTION (provided by applicant): Diseases caused by Trypanosoma brucei spp. represent health hazards for many disadvantaged countries. Because of failing vector control and the rise of drug-resistance, the re-appearance of African sleeping sickness necessitates a search for new drug targets with emphasis on parasite-specific processes. Trypanosomal mitochondrial RNA processing pathways are extremely divergent from those of humans and represent a potential source of therapeutic intervention points. The mRNA editing has been extensively studied, but the significance of pre- and post-editing processing events emerged only recently. We discovered the mitochondrial poly (A) polymerase (KPAP1) and terminal uridyltransferase (RET1), and showed that intertwined polyadenylation and uridylation processes are critical for mRNA stability, translation and decay. In these studies, I encountered a diverse family of pentatricopeptide (PPR, 35 amino acids) helical repeat- containing RNA binding proteins that populate polyadenylation and translation complexes. By focusing on mRNA 3' end definition, modification and stability, this proposal introduces PPR proteins as key regulators of mitochondrial gene expression. The experimental approaches involve biochemistry, proteomics, crystallography, genetics and deep RNA sequencing. The proposal aims to: 1) Dissect general and transcript-specific mRNA adenylation/uridylation mechanisms. I hypothesize that the two modes of 3' modification, pre-editing addition of a short A-tail and post-editing A/U-tailing, serve discrete purposes of mRNA stabilization and commitment to translation, respectively. 2) Determine principles of uridylation- based mRNA decay. A hypothesis that RET1 TUTase-catalyzed uridylation accelerates mRNA 3'-5' degradation will be tested. I also posit that most transcripts are stabilized by a specific PPR protein that blocks mRNA uridylation. 3) Elucidate functions of a putative mRNA 3' definition and stabilization factor. I suggest that a PPR factor is deposited onto the pre-mRNA prior to polyadenylation to define the 3' end. We focus on select PPR proteins in trypanosomes, but the long term goal is to illuminate the mechanisms of RNA recognition by repeat-containing proteins, which will be of fundamental importance for RNA processing and mitochondrial biology fields.

 描述（由适用提供）：由Brucei spp引起的疾病。代表许多弱势国家的健康危害。由于载体的控制失败和抗药性的兴起，新药靶标需要的非洲睡眠疾病的重新表现，重点是寄生虫特异性过程。锥虫线粒体RNA处理途径与人类的锥虫非常多样化，代表了热干预点的潜在来源。 mRNA编辑已经进行了广泛的研究，但是在最近才出现了预编辑事件的重要性。我们发现了线粒体聚（A）聚合酶（KPAP1）和末端尿苷转移酶（RET1），并表明相互论形的聚腺苷酸化和尿苷过程对于mRNA稳定性，翻译和衰减至关重要。在这些研究中，我遇到了五肽的潜水家族（PPR，35个氨基酸）螺旋重复 - 含有填充聚腺苷酸化和翻译复合物的RNA结合蛋白。通过关注mRNA 3'末端定义，修饰和稳定性，该建议将PPR蛋白作为线粒体基因表达的关键调节剂。实验方法涉及生物化学，蛋白质组学，晶体学，遗传学和深度RNA测序。该提案的目的是：1）剖析一般和转录特定的mRNA腺苷酸化/尿苷机制。我假设3'修改的两种模式，预编辑简短的A尾和编辑后A/U-tailing，分别实现了mRNA稳定和对翻译的承诺的离散目的。 2）确定基于尿苷的mRNA衰减的原理。将测试一个假说，即ret1诱导酶催化的尿苷加速了mRNA 3'-5'降解。我还认为，大多数转录本都通过阻断mRNA尿素化的特定PPR蛋白稳定。 3）阐明假定mRNA 3'定义和稳定因子的功能。我建议将PPR因子沉积到聚腺苷酸化之前的前mRNA上，以定义3'端。我们专注于锥虫中的某些PPR蛋白，但长期目标是通过重复含有蛋白质来阐明RNA识别的机制，这对于RNA加工和线粒体生物学领域至关重要。