Role of a novel PGC-1α isoform in gene transcription

新型 PGC-1α 亚型在基因转录中的作用

• 批准号: 10525004
• 负责人: Xavier Rambout
• 金额: $ 19.25万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10525004
• 关键词: Adhesions; Age; Aging; Alleles; Amino Acid Sequence; Amino Acids; Anti-Inflammatory Agents; Antibodies; Antisense Oligonucleotides; Area; Atherosclerosis; Binding; Biological Assay; Blood Vessels; C-terminal; CCL2 gene; CCL7 gene; CCL8 gene; CRISPR/Cas technology; Cardiac; Cardiac development; Cardiovascular Diseases; Cause of Death; Cellular Metabolic Process; Chemotactic Factors; Chromatin; Complementary DNA; Data; Development; Disease; Dystrophin; Endothelial Cells; Endothelium; Engineering; Exons; Female; Gene Expression Regulation; Genes; Genetic Transcription; In Vitro; Individual; Infiltration; Inflammation; Inflammatory; Inflammatory Response; Knock-out; Liquid Chromatography; MAP Kinase Gene; Malignant Neoplasms; Mammalian Cell; Maps; Messenger RNA; Metabolic; Metabolism; Modeling; Monocyte Chemoattractant Proteins; Mus; Muscle; Mutate; Myoblasts; Names; Nucleotides; PPAR gamma; Phenocopy; Plasma; Play; Polymerase Chain Reaction; Premature aging syndrome; Production; Protein Isoforms; Proteins; RNA; RNA Cap-Binding Proteins; RNA Polymerase II; RNA Splicing; RNA-Binding Proteins; Regulation; Reporting; Research; Research Personnel; Resolution; Risk; Risk Factors; Role; Running; Signal Transduction; Skeletal Muscle; Small Interfering RNA; Stress; Structure; Testing; Tissues; Transactivation; Transcript; Transcription Coactivator; Transcriptional Regulation; Variant; alpha helix; base; career; chemokine; coronary fibrosis; cytokine; design; functional loss; heart function; in vivo; knock-down; mRNA Precursor; male; migration; monocyte; mouse model; novel; novel therapeutics; physiologic stressor; promoter; recruit; response; sex; systemic inflammatory response; tandem mass spectrometry; transcription factor; transcriptome sequencing

Peroxisome proliferator-activated receptor gamma co-activator 1α (PGC-1α) is a transcriptional coactivator that is critical for the metabolic adaptation of mammalian cells to diverse physiological stresses. In addition to this well-described function in the control of cell metabolism, the number of reports describing a protective role for PGC-1α against inflammation is on the rise. I report here a novel stress-dependent pro-inflammatory PGC-1α variant that I have identified in vitro in C2C12 mouse myoblasts and in vivo in mouse skeletal muscle. My unpublished results indicate that, compared to the canonical PGC-1α isoform (also known as PGC-1α(1)), the structure of this novel 72-kDa PGC-1α isoform, which I will call PGC-1α(72), is typified by its lack of the C-terminal 15 amino acids. The deleted C-terminal sequence encompasses a 9-amino-acid alpha helix that I previously reported to be essential for PGC-1α(1) to bind to the cap-binding protein (CBP)80 and to transactivate pro-myogenic PGC-1α-dependent genes. In contrast to PGC-1α(1), PGC-1α(72) lacks the CBP80-binding motif (CBM) and, as my data indicate, activates the transcription of pro-inflammatory genes. In the first aim of my research strategy, I propose to use C2C12 myoblasts to fully characterize the sequence of this novel PGC-1α(72) isoform, identify those transcription factors and RNA-binding proteins that regulate its expression in response to stress, and comprehensively determine the transcriptional network that it activates. In the second aim, I will take advantage of my finding that functional inhibition of the PGC-1α CBM phenocopies PGC-1α(72)-dependent expression of pro-inflammatory chemoattractants such as the chemokine CCL2. Elevated CCL2 plasma levels, which are observed with aging, have been recognized as important risk factors for numerous inflammatory diseases, including cardiac fibrosis, atherosclerosis and cancers. Mechanistically, CCL2 has been found to function in monocyte recruitment onto the endothelium of blood vessels and in monocyte infiltration into inflamed tissues. I will breed our newly characterized PGC-1αCBMmut mice with the Dystrophin-deficient Dmdmdx muscle inflammatory and premature aging mouse model and use derived primary myoblasts to infer the functional consequences of PGC-1α(72) expression, in particular those attributable to its lacking the CBM, to inflammation. In particular, I will determine how PGC-1αCBMmut triggers the CCL2-dependent recruitment of monocytes onto endothelial cells, their infiltration into cardiac tissues, and development of cardiac fibrosis in aging mice as a function of sex, i.e. in males vs. females. I view demonstrating the contribution of PGC-1αCBMmut to cardiac fibrosis as a proof of concept. It will provide fertile grounds to study the function of skeletal muscle PGC-1α(72) in the development of other aging- related inflammatory diseases. I hypothesize that understanding PGC-1α(72)-dependent gene regulation and its consequences to the development of inflammatory diseases will facilitate the development of new therapeutics, such as synthetic antisense oligonucleotides regulating splicing of PGC-1α pre-mRNA.

过氧化物体增生剂激活受体伽马共激活剂1α（PGC-1α）是转录共激活因子， 对于哺乳动物细胞对潜水身体应激的代谢适应至关重要。除此之外 在控制细胞代谢的控制方面有很好的描述功能，描述了保护作用的报告数量 PGC-1α反对炎症正在上升。我在这里报告了一种新型的应力依赖性促炎PGC-1α 我在C2C12小鼠成肌细胞和小鼠骨骼肌中体内鉴定的变体。 我未发表的结果表明，与规范的PGC-1α同工型相比 PGC-1α（1）），这款新型72-kDa PGC-1α同工型的结构，我称为PGC-1α（72），是其典型的 缺乏C末端15个氨基酸。已删除的C末端序列包括9-氨基酸α 我以前据报道的螺旋对于PGC-1α（1）与结合帽结合蛋白（CBP）80和对 反式激活促肌PGC-1α依赖性基因。与PGC-1α（1）相反，PGC-1α（72）缺乏 CBP80结合基序（CBM），作为我的数据指标，激活了促炎基因的转录。在 我的研究策略的第一个目的，我建议使用C2C12成肌细胞充分表征 这种新型的PGC-1α（72）同工型，鉴定那些转录因子和调节其调节的RNA结合蛋白 响应压力的表达，并全面确定其激活的转录网络。 在第二个目标中，我将利用发现PGC-1αCBM的功能抑制 表型PGC-1α（72） - 依赖性促趋化趋化剂（例如趋化因子）的表达 CCL2。通过衰老观察到的升高的CCL2血浆水平已被认为是重要的风险 许多炎症性疾病的因素，包括心脏纤维化，动脉粥样硬化和癌症。 从机械上讲，已经发现CCL2在单核细胞募集到血液内皮中起作用 血管和单核细胞浸润到发炎的组织中。我将繁殖我们新特征的PGC-1αCBMMUT 具有肌营养不良蛋白的DMDMDX肌肉炎症和过早老化小鼠模型的小鼠并使用 衍生的原代肌细胞来推断PGC-1α（72）表达的功能后果，特别是 归因于其缺乏CBM的炎症。特别是，我将确定PGC-1αCBMMUT如何触发 CCL2依赖性单核细胞募集到内皮细胞上，它们浸润到心脏组织中，并 衰老小鼠的心脏纤维化发展是性别的作用，即男性与女性。 我认为证明PGC-1αCBMMUT对心脏纤维化的贡献是概念证明。会 提供肥沃的理由来研究骨骼肌PGC-1α（72）在其他衰老发展中的功能 相关炎症性疾病。我假设了解PGC-1α（72）依赖性基因调节及其 炎症性疾病发展的后果将促进新治疗学的发展， 例如合成反义寡核苷酸调节PGC-1α前MRNA的剪接。