Structure to phenotype analysis of a conserved RNA binding protein required for reproduction

繁殖所需的保守 RNA 结合蛋白的结构到表型分析

基本信息

批准号：
10478905
负责人：
Francesca Massi
金额：
$ 51.22万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10478905
关键词：
Affect Amino Acid Sequence Amino Acids Amyotrophic Lateral Sclerosis Animals Anterior Binding Biochemical Biochemistry Biological Biological Process CRISPR/Cas technology Caenorhabditis elegans Cell division Cells Child Computer Simulation Cytoplasm Data Defect Development Embryo Embryonic Development Event Family Family member Fertility Fertilization Gene Activation Gene Transfer Techniques Goals In Vitro Libraries Malignant Neoplasms Maps Maternal Messenger RNA Messenger RNA Methods Molecular Mothers Mutagenesis Mutate Mutation Myotonic Dystrophy NMR Spectroscopy Oocytes Oogenesis Orthologous Gene Pathway interactions Pattern Phenotype Post-Transcriptional Regulation Process Property Protein Dynamics Protein Family Proteins Publishing RNA RNA Binding RNA Sequences RNA Splicing RNA-Binding Proteins Regulation Relaxation Reporter Repression Reproduction Reproductive Biology Research Role Site-Directed Mutagenesis Specificity Structure Testing Transcript Translations Work Zinc Fingers biophysical techniques cell fate specification cell type developmental disease experimental study genome editing human disease in vivo innovation insight molecular dynamics oocyte maturation polarized cell prevent protein structure segregation

项目摘要

During metazoan development, polarization of the body axes is of critical importance, as asymmetric cell division initiates cell specialization pathways. A family of conserved RNA-binding proteins characterized by CCCH-type tandem zinc finger (TZF) domains is required for axis polarization and cell type specification in the early embryo of C. elegans. A cascade of events essential to the oocyte- to-embryo transition is initiated by two members of this family, OMA-1/2. The process initiates with OMA-1/2 turnover and culminates with asymmetric segregation of cellular components to opposing poles of the embryo. Mutation of OMA-1/2 blocks oocyte maturation and prevents fertilization. We have observed that OMA-1 binds RNA with relatively low specificity in vitro, but regulates specific mRNAs in vivo. We have also determined that OMA-1 RNA binding is highly cooperative, a feature never before observed in a protein from the CCCH-type TZF family. Our goal is to investigate OMA- 1/2 activity from structure to phenotype. We will determine the molecular mechanism or RNA-binding cooperativity using NMR spectroscopy, computer simulations, and quantitative biochemistry. We will also define how RNA-binding cooperativity defines OMA-1 biological activity in worms. We will determine which mRNAs are regulated by OMA-1/2 in oocytes and of these mRNAs which are the most critical to reproduction. This research will lead to a new understanding of how the molecular properties of OMA-1 determine its function in reproductive biology in living animals.

在多宗发育过程中，人体轴的极化至关重要，因为不对称细胞分裂启动细胞专业途径。保守的RNA结合蛋白家族轴极化和秀丽隐杆线虫早期胚胎中的细胞类型规格。卵母细胞必不可少的事件级联该家族的两个成员OMA-1/2发起了待办事项过渡。该过程启动 OMA-1/2周转率，并以对相对的细胞成分的不对称分离达到顶点胚胎的杆。 OMA-1/2的突变阻断卵母细胞的成熟并防止受精。我们已经观察到OMA-1在体外具有相对较低的特异性结合RNA，但调节特定 MRNA在体内。我们还确定OMA-1 RNA结合是高度合作的，这是一个特征从CCCH型TZF家族的蛋白质中从未观察到。我们的目标是调查OMA- 从结构到表型的1/2活性。我们将确定分子机制或RNA结合使用NMR光谱，计算机模拟和定量生物化学的合作性。我们将还定义了RNA结合合作性如何定义蠕虫中的OMA-1生物学活性。我们将确定哪些mRNA在卵母细胞中受OMA-1/2的调节，而这些mRNA是最重要的是生殖。这项研究将导致对分子的新了解 OMA-1的特性决定了其在活动物中生殖生物学中的功能。