权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

RNA Processing Machines in Biology and Disease

生物学和疾病中的 RNA 加工机器

基本信息

批准号：
10605338
负责人：
DANESH MOAZED
金额：
$ 19.15万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-04-01 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10605338
关键词：
Affect Amyotrophic Lateral Sclerosis Antigen Presentation Antisense Technology Biological Assay Biology Cells Central Nervous System Clustered Regularly Interspaced Short Palindromic Repeats Coculture Techniques DNA Binding DNA-Binding Proteins Defect Disease Dysmyelopoietic Syndromes Gene Expression Genes Goals Hematologic Neoplasms Hematopoiesis Hematopoietic Hematopoietic Neoplasms Human Biology Immune Immune system Malignant Neoplasms Microglia Motor Neuron Disease Motor Neurons Mutate Mutation Pathogenesis Play Proteins Proteomics RNA RNA Processing RNA Splicing RNA, Messenger, Splicing Research Research Project Grants Role Spliceosomes System TAF15 gene Technology Therapeutic Transcriptional Regulation Work causal variant embryonic stem cell genetic signature hematopoietic differentiation human disease human embryonic stem cell mRNA Precursor mutant neuron loss new therapeutic target targeted treatment transcriptomics

项目摘要

PROJECT SUMMARY / ABSTRACT The goal of this work is to understand the disease-causative roles and basic biology of the human spliceosome. Pre-mRNA splicing is the best-known function of the spliceosome, but this machinery also houses RNA/DNA binding proteins with roles in many steps of gene expression. The lab focuses on the motor neuron disease amyotrophic lateral sclerosis (ALS) and on blood cancers. Greater than one third of ALS-causative genes encode RNA/DNA binding proteins yet their functions are not well understood. Our new research led to the exciting discovery that three of these proteins (FUS, TAF15, MATR3) are essential for expression of a set of antigen presentation genes, which play critical protective roles in the immune system. Remarkably, the three ALS genes are also required for expression of the master transcription control factor of the antigen presentation genes. In ALS, hyperactivation of the immune system is known for its detrimental effects on motor neurons. However, the immune system is emerging as a double-edged sword in ALS as studies indicate that loss of its protective functions also contributes to motor neuron death. At present, little is known about the protective roles. To determine whether loss of the antigen presentation factors due to mutant ALS genes contributes to ALS, human embryonic stem (ES) cells will be CRISPR-edited to harbor ALS-causative mutations in the RNA/DNA binding genes. The ES cells will be differentiated into microglia, which are the immune cells of the central nervous system. Co-culture systems will be used to determine how motor neurons are affected by the mutant microglia. Proteomics, transcriptomics, and functional assays will be used to assess effects on microglia and motor neurons. A critical objective of the work is to determine whether loss of the antigen presentation factors can be extended to other forms of ALS. If so, it raises the exciting possibility that therapies targeting these factors may be efficacious for multiple types of ALS. The goal of our other research project is to determine how mutation of the spliceosomal protein SF3B1 contributes to blood cancers. We CRISPR-edited ES cells to harbor an SF3B1 cancer mutation and will differentiate the ES cells into hematopoietic lineages. Transcriptomics/proteomics and impacts on differentiation will be used to identify changes that may contribute to SF3B1 cancers. We identified a set of mis-splicing signature genes common to different SF3B1 cancers and will determine how their mis- splicing affects differentiation. Mis-splicing of signatures that are candidates for contributing to SF3B1 cancers will be corrected using antisense technology, with the goal of developing the technology as a therapeutic. Finally, we identified two genes that are robustly upregulated only in spliceosome-mutated myelodysplastic syndrome (MDS), which is main type of blood cancer associated with spliceosome mutations. Both upregulated genes play key roles in hematopoiesis. Their potential contribution to MDS will be examined using the hematopoietic differentiation assays. Together, this research will lead to important advances in identifying the roles of defective spliceosomal genes in both ALS and hematological cancers as well as aid in identifying new therapeutic targets.

项目概要/摘要这项工作的目标是了解人类剪接体的致病作用和基本生物学。 mRNA 前体剪接是剪接体最著名的功能，但该机器还包含 RNA/DNA 结合蛋白在基因表达的许多步骤中发挥作用。该实验室专注于运动神经元疾病肌萎缩侧索硬化症（ALS）和血癌。超过三分之一的 ALS 致病基因编码 RNA/DNA 结合蛋白但其功能尚不清楚。我们的新研究带来了令人兴奋的结果发现其中三种蛋白质（FUS、TAF15、MATR3）对于一组抗原的表达至关重要呈递基因，在免疫系统中发挥重要的保护作用。值得注意的是，三个 ALS 基因抗原呈递基因的主转录控制因子的表达也需要这些。在 ALS（免疫系统过度激活）因其对运动神经元的有害影响而闻名。然而，免疫系统正在成为治疗 ALS 的一把双刃剑，因为研究表明，免疫系统的保护作用丧失功能也会导致运动神经元死亡。目前，人们对其保护作用知之甚少。到确定由于 ALS 基因突变导致的抗原呈递因子的丧失是否会导致 ALS，人类胚胎干 (ES) 细胞将经过 CRISPR 编辑，以在 RNA/DNA 结合中隐藏 ALS 致病突变基因。 ES细胞将分化为小胶质细胞，小胶质细胞是中枢神经的免疫细胞系统。共培养系统将用于确定突变小胶质细胞如何影响运动神经元。蛋白质组学、转录组学和功能测定将用于评估对小胶质细胞和运动神经元的影响神经元。这项工作的一个关键目标是确定抗原呈递因子的丧失是否可以扩展到其他形式的 ALS。如果是这样，那就提出了一种令人兴奋的可能性，即针对这些因素的疗法可能会对多种类型的 ALS 有效。我们另一个研究项目的目标是确定突变是如何发生的剪接体蛋白 SF3B1 会导致血癌。我们对 ES 细胞进行 CRISPR 编辑，使其含有 SF3B1 癌症突变并将 ES 细胞分化为造血谱系。转录组学/蛋白质组学和对分化的影响将用于识别可能导致 SF3B1 癌症的变化。我们确定了一组不同 SF3B1 癌症常见的错误剪接特征基因，将决定它们如何错误剪接剪接影响分化。导致 SF3B1 癌症的候选签名错误拼接将使用反义技术进行纠正，目标是将该技术开发为治疗方法。最后，我们发现了两个仅在剪接体突变的骨髓增生异常综合征中显着上调的基因（MDS），这是与剪接体突变相关的主要血癌类型。两个上调基因都发挥作用在造血中发挥关键作用。将使用造血系统检查它们对 MDS 的潜在贡献分化分析。总之，这项研究将在识别缺陷的作用方面取得重要进展。 ALS 和血液癌症中的剪接体基因，并有助于确定新的治疗靶点。