Erythroid stage-specific transcriptome expression, dynamics, and regulation

红系阶段特异性转录组表达、动态和调控

基本信息

批准号：
8258173
负责人：
JOHN G CONBOY
金额：
$ 45.14万
依托单位：
UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-20 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8258173
关键词：
Address Affect Alternative Splicing Basophilic Erythroblast Binding Biochemical Bioinformatics Biological Models Biological Process Biology Bone Marrow Cell division Cell physiology Cells Characteristics Complex Computer Analysis Cultured Cells Cytoskeleton Data Data Analyses Defect Detection Development Ensure Erythroblasts Erythrocytes Erythroid Erythroid Cells Erythropoiesis Event Evolution Exhibits Exons Fetal Liver Foundations Future Gene Expression Profile Genome Gifts Goals Health Hereditary Disease Human Human Biology Human Genetics Introns Joints Knockout Mice Knowledge Lead Length Methods Mining Modeling Mus Myoblasts Nucleotides Oligonucleotides Pathway interactions Pattern Physiological Population Post-Transcriptional Regulation Pronormoblasts Property Protein Isoforms Proteins Proteome RNA RNA Processing RNA Splicing RNA analysis Regulation Regulatory Element Research Research Personnel Shapes Sorting - Cell Movement Splicing Regulation Pathway Staging Structure System Systems Analysis Testing Tissues Transcript Transcriptional Regulation Work base chromatin modification comparative genomics erythroid differentiation expectation genetic regulatory protein human disease improved innovation insight iron metabolism mRNA Precursor novel programs response tool

项目摘要

DESCRIPTION (provided by applicant): Primary focus will be on global characterization of late erythroid transcriptome(s) with emphasis on detection of new transcripts and new isoforms of known transcripts, dynamic evolution of the transcriptome during late erythropoiesis, and mechanisms by which an evolutionarily conserved alternative splicing program shapes the transcriptome. Differentiating erythroblasts execute a diverse and dynamic pre-mRNA alternative splicing program that cooperates with the transcriptional program to ensure synthesis of the appropriate stage-specific proteome as cells acquire specialized functional properties. Proper regulation of alternative splicing is extremely relevant to human health, for misregulation is a major contributor to many human diseases yet the erythroid splicing program, its regulation, and its importance in erythroid biology remain poorly understood. This project proposes a global analysis of the stage-specific erythroid transcriptome by RNA-Seq analysis and advanced bioinformatic strategies to address these issues and to generate a wealth of new information of use to many other investigators studying erythroid differentiation and erythroid biology. To explore the hypothesis that a conserved mammalian erythroid alternative splicing program regulates critical erythroid functions, investigators with expertise in erythroid differentiation, alternative splicing regulation, and computational analysis of deep sequencing data have come together to propose three specific research aims. Aim 1 will define the mouse erythroid stage-specific transcriptome using highly purified FACS-sorted erythroid cells from bone marrow (proerythroblasts as well as basophilic, polychromatic, and orthochromatic erythroblast stages). Advanced computational analysis of RNA-seq data enables comparison among the differentiation stages and between erythroid and non-erythroid cells, to characterize erythroid isoform diversity and stage-specific switches in alternative splicing that imply functional changes in the encoded proteome. Aim 2 will perform a similar analysis of human erythroblasts that are highly purified by FACS sorting. Comparison of human and mouse data will facilitate the definition of evolutionarily conserved erythroid-specific and dynamic switches in isoform expression that suggests critical erythroid functions, in addition to highlighting isoform differences that exist between mouse and human cells. Aim 3 proposes a mechanistic study of the conserved alternative splicing events defined in Aims 1 and 2 by using computational and biochemical approaches to analyze cis- regulatory sequences and splicing factor proteins that direct these splicing networks. Ultimately, this work should reveal the splicing regulatory network(s) that orchestrate programmed splicing in differentiating erythroid cells. Long term benefits anticipated from this work include greatly improved insights into regulation of biological processes in erythroid cells by alternative protein isoforms. Moreover, the RNA-Seq data itself may stimulate studies of the transcriptional and post-transcriptional regulation of this transcriptome. PUBLIC HEALTH RELEVANCE: Understanding how tissue-specific alternative splicing is regulated is extremely relevant to human health, because splicing mis-regulation is the underlying cause of a significant number of human genetic diseases. This project will lay the foundations for analysis of alternative splicing networks in that operate during differentiation of erythroblasts into mature red cells. This knowledge will stimulate mechanistic studies of normal red cell function and may contribute to understanding of aberrant red cells due to splicing defects.

描述（由申请人提供）：主要重点将放在晚期红细胞转录组的全球表征上，重点是检测新的转录本已知转录物的新转录物和新的同工型，eriythropoiesis晚期转录组的动态演化，以及该机制的机制以及该进化保守的交流型剪接程序形状的转录组。区分红细胞，执行多种多样的动态前MRNA替代剪接程序，该程序与转录程序合作，以确保将适当的阶段特异性蛋白质组合成，因为细胞获得了专门的功能性能。对替代剪接的适当调节与人类健康非常相关，因为误导是许多人类疾病的主要因素，但红斑剪接计划，调节及其在红细胞生物学中的重要性仍然很众所周知。该项目提出了通过RNA-Seq分析和先进的生物信息学策略对特定阶段的红细胞转录组进行全球分析，以解决这些问题，并向许多其他研究红细胞分化和红细胞生物学的研究者生成大量新的使用信息。为了探讨以下假设：保守的哺乳动物红斑替代剪接程序调节关键的红斑功能，研究人员在红细胞分化方面具有专业知识，替代剪接调控以及深层测序数据的计算分析，以提出三个特定的研究目的。 AIM 1将使用高度纯化的FACS分类的红细胞细胞来定义小鼠粒子阶段特异性转录组（骨髓以及嗜碱性，嗜碱性，多色和正色成红细胞阶段）。 RNA-seq数据的高级计算分析可以比较分化阶段以及红细胞和非果皮细胞之间的比较，以表征替代剪接中的红斑同工型多样性和特定于阶段的转换，这意味着编码蛋白质组的功能变化。 AIM 2将对FACS分类高度纯化的人类红细胞进行类似的分析。人类和小鼠数据的比较将促进同工型表达中进化保守的红细胞特异性和动态开关的定义，除了突出了小鼠和人类细胞之间存在的同工型差异外，还表明了关键的红细胞功能。 AIM 3提出了通过使用计算和生化方法来分析指导这些剪接网络的顺式调节序列和剪接因子蛋白，对目标1和2中定义的保守替代剪接事件的机械研究。最终，这项工作应揭示剪接调节网络，该网络在区分红细胞中编程的拼接。这项工作预期的长期益处包括通过替代蛋白质同工型大大提高了对红细胞细胞生物过程调节的见解。此外，RNA-seq数据本身可能会刺激该转录组的转录和转录后调节的研究。公共卫生相关性：了解组织特异性替代剪接的调节与人类健康极为相关，因为剪接错误调节是大量人类遗传疾病的根本原因。该项目将奠定基础，用于分析替代剪接网络，该网络在将红细胞分化为成熟的红细胞时运行。这些知识将刺激正常红细胞功能的机理研究，并可能有助于理解由于剪接缺陷而引起的异常红细胞。