FOXO signaling and skeletal muscle atrophy

FOXO 信号传导与骨骼肌萎缩

基本信息

批准号：
8676431
负责人：
Andrew Robert Judge
金额：
$ 31.71万
依托单位：
UNIVERSITY OF FLORIDA
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：
2011-08-01 至 2016-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8676431
关键词：
Acetylation Acetyltransferase Acquired Immunodeficiency Syndrome Address Aging Area Atrophic Bed rest Binding Boxing CREB-binding protein Cells Chronic Chronic Obstructive Airway Disease DNA Binding Data Deacetylase Deacetylation Denervation Diabetes Mellitus Disease Dominant-Negative Mutation EP300 gene Electrophoretic Mobility Shift Assay Family Gene Targeting Genetic Genetic Transcription Goals HDAC2 gene HDAC4 gene Health Heart failure IGF1 gene Immobilization In Vitro Intervention Lead MLLT7 gene Malignant Neoplasms Measures Mediating Muscle Muscle Fibers Muscular Atrophy Nuclear Nutrient Oligonucleotides Pathway interactions Phosphorylation Physiological Plasmids Post-Translational Protein Processing Protein Acetylation Protein Family Proteins Regulation Reporter Research Role Sepsis Signal Pathway Signal Transduction Signaling Molecule Skeletal Muscle Starvation Stimulus Testing Transcriptional Activation Transfection Work base cell type deprivation human CREBBP protein improved in vivo mRNA Expression muscle form mutant p300/CBP-Associated Factor programs research study response skeletal muscle wasting therapeutic target tool transcription factor wasting

项目摘要

DESCRIPTION (provided by applicant): Skeletal muscle atrophy is a widespread physiological phenomenon and significant health problem associated with muscle disuse (immobilization, bedrest, denervation) and various diseases (cancer, sepsis, AIDS, diabetes, chronic heart failure, chronic obstructive pulmonary disease). However, our understanding of the signaling molecules that regulate muscle mass during an atrophy condition are ill defined. Therefore the long- range goal of our research program is to understand the regulation of signaling pathways that cause muscle atrophy during various conditions. Eventually improved understanding will lead to the identification of the most suitable targets for specific interventions. One family of proteins that is known to cause skeletal muscle atrophy is the FOXO family, and FOXO signaling is increased in a variety of atrophy conditions, including cast immobilization, sepsis, cancer and starvation. However, our understanding of the regulation of FOXO signaling in skeletal muscle is largely confined to the phosphorylation of FOXO factors via the IGF1/PI3K/Akt pathway. While this pathway is clearly important in the regulation of FOXO signaling, another mechanism of regulation, acetylation, appears to be equally important in regulating FOXO signaling, at least in cultured non-muscle cell. However, it is currently unknown whether acetylation of FOXO factors is a regulatory mechanism of FOXO signaling in skeletal muscle during any condition of muscle atrophy. The overall objective of the current proposal is to determine the role of acetyltransferase (HAT) proteins and deacetylase (HDAC) proteins, and acetylation and deacetylation of FOXO factors, in regulating FOXO signaling as it relates to skeletal muscle atrophy. To address this, in Aim 1 we will transfect C2C12 skeletal muscle cells with wild type (WT) or dominant negative (d.n.) HAT proteins and determine their regulation of endogenous FOXO signaling. We will subsequently select one HAT protein for further study in whole skeletal muscle during muscle disuse and cancer. In Aim 2 we will transfect C2C12 skeletal muscle cells with wild type (WT) or dominant negative (d.n.) HDAC proteins and determine their regulation of endogenous FOXO signaling. We will subsequently select one HDAC protein for further study in whole skeletal muscle during muscle disuse and cancer. In Aim 3 we will transfect C2C12 skeletal muscle cells with plasmids encoding FOXO acetylation mutants that mimic either the acetylated or deacetylated forms of FOXO and determine their regulation of FOXO signaling. In each of these Aims we will measure FOXO transcriptional activity, FOXO- DNA binding, FOXO cellular localization, post-translational modifications of FOXO factors, the transcription of a sub-set of known FOXO target genes, and skeletal muscle fiber cross sectional area. The findings from these experiments will lead to a greater understanding of the regulation of FOXO signaling as it relates to skeletal muscle atrophy.

描述（由申请人提供）：骨骼肌萎缩是一种广泛的生理现象，与肌肉残疾相关的重大健康问题（固定，床，卧室，脱腐）和各种疾病（癌症，脓毒症，艾滋病，艾滋病，糖尿病，糖尿病，慢性心力衰竭，慢性阻塞性肺疾病）。但是，我们对调节萎缩状况下调节肌肉质量的信号传导分子的理解是不明的。因此，我们的研究计划的长距离目标是了解在各种情况下引起肌肉萎缩的信号通路的调节。最终，改善的理解将导致确定特定干预措施的最合适目标。 FOXO家族是众所周知的一个引起骨骼肌萎缩的蛋白质家族，在多种萎缩条件下，Foxo信号传导增加，包括铸造固定，败血症，癌症和饥饿。但是，我们对骨骼肌中FOXO信号的调节的理解在很大程度上仅限于通过IGF1/PI3K/AKT途径的FoxO因子的磷酸化。尽管这种途径在调节FOXO信号传导中显然很重要，但至少在培养的非肌肉细胞中，调节的另一种调节机制，乙酰化似乎同样重要。但是，目前尚不清楚FOXO因子的乙酰化是否是肌肉萎缩任何条件下骨骼肌中FOXO信号传导的调节机制。当前建议的总体目的是确定乙酰转移酶（HAT）蛋白和脱乙酰基酶（HDAC）蛋白的作用，以及FOXO因子的乙酰化和脱乙酰基在调节与骨骼肌萎缩有关的FOXO信号传导中的作用。为了解决这个问题，在AIM 1中，我们将使用野生型（WT）或显性负（D.N.）HAT蛋白转染C2C12骨骼肌细胞，并确定其调节内源性FOXO信号传导。随后，我们将在肌肉消失和癌症期间选择一种帽子蛋白，以进一步研究整个骨骼肌。在AIM 2中，我们将使用野生型（WT）或显性阴性（D.N.）HDAC蛋白转染C2C12骨骼肌细胞，并确定其对内源性FOXO信号传导的调节。随后，我们将选择一种HDAC蛋白，以在肌肉消失和癌症期间进一步研究整个骨骼肌。在AIM 3中，我们将用编码FOXO乙酰化突变体的质粒转染C2C12骨骼肌细胞，这些突变体模仿FoxO的乙酰化或脱乙酰化形式，并确定其调节FOXO信号传导。在这些目标中，我们将测量FOXO转录活性，Foxo-DNA结合，FOXO细胞定位，FOXO因子的翻译后修饰，已知FOXO靶基因的子集的转录以及骨骼肌纤维纤维横截面区域。这些实验的发现将导致对FOXO信号传导的调节，因为它与骨骼肌萎缩有关。