Molecular regulation of adipocyte progenitor quiescence and metabolic adaptation to obesity

脂肪细胞祖细胞静止的分子调控和肥胖代谢适应

基本信息

批准号：
10419976
负责人：
Matthew Steinhauser
金额：
$ 25.62万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-05-17 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10419976
关键词：
ATAC-seq Address Adipocytes Adipose tissue Adult Affect Binding Biological Assay CRISPR interference CRISPR/Cas technology Calories Cells Chromatin Coupled Coupling DNA DNA Binding Domain Data Development Developmental Gene Diabetes Mellitus Enhancers Failure Familial generalized lipodystrophy Fatty acid glycerol esters Future Genes Genetic Transcription Genetic Variation Genomics Growth Health Heterogeneity Hyperplasia Hypertrophy Impairment In Vitro Insulin Resistance Knowledge Life Link Maps Mass Spectrum Analysis Metabolic Methodology Methods Modeling Molecular Morbidity - disease rate Mus NR4A1 gene Non-Insulin-Dependent Diabetes Mellitus Obesity Output PPARG gene Pathway interactions Population Proliferating Proxy Publishing Regulation Reporter Risk Role Sorting - Cell Movement Source Structure Testing Tissue Expansion Tissues Tracer Work adipocyte differentiation age related experimental study functional genomics genetic variant genome-wide in vivo lipid biosynthesis loss of function mortality network models prevent progenitor programs response stable isotope stem cells transcription factor transcriptomics

项目摘要

Adipose tissue expansion occurs by a combination of new adipocyte formation (resulting in hyperplastic growth) and hypertrophy of existing adipocytes, the relative contributions of which may impact systemic metabolic health. Congenital lipodystrophy due to failed adipocyte development is associated with severe insulin resistance. More subtle impairments in adipogenesis arising from genetic variants in developmental genes also predispose to insulin resistance and Type 2 Diabetes Mellitus. These examples reflect a larger body of data providing a conceptual basis for the hypothesis that metabolic health is dependent on a functional adipocyte population. By leveraging stable isotope tracers with high precision mass spectrometry methods, we have now also identified an age-dependent decline in adipogenesis. Therefore, adipocyte progenitors—the cellular source of new adipocytes—must reside in a state of relative quiescence in mature adipose tissue. Although downstream drivers of adipogenesis are well described, the mechanisms that maintain quiescence of adipocyte progenitors or their proximal transition to proliferate and differentiate, in vivo, remain largely unknown. To address this knowledge gap, we have leveraged transcriptomic data together with genome scale maps of accessible chromatin in freshly isolated primary adipocyte progenitors to model the network of transcription factors responsible for the quiescent state. Our data implicate the orphan nuclear receptor, NR4A1, as a key regulatory node. In vitro and in vivo functional genomics studies have provided additional support for our NR4A1-centric hypothesis: that NR4A1 regulates a transcriptional program that establishes a metabolically deleterious state of quiescence in adipocyte progenitors. These data also provide rationale for two interrelated Specific Aims. In Aim 1, we will test the hypothesis that NR4A1 is a core transcriptional regulator of adipocyte progenitor quiescence. We will use reporter assays, CRISPRi and CRISPR/Cas9 to functionally interrogate NR4A1-DNA regulatory nodes at key adipogenic transcription factors. In Aim 2, we will perform an unbiased interrogation of the NR4A1-depedent chromatin regulatory landscape in adipocyte progenitor cells at genome scale. We will map NR4A1-DNA interactions and co-localize NR4A1 binding with canonical cis-regulatory marks. We will then use low-input genomics methods coupled with NR4A1 gain or loss of function to map the NR4A1 dependent chromatin regulatory landscape and transcriptional output. Collectively, through targeted functional genomics experiments and unbiased genome scale analyses we will test our model of NR4A1 as a master regulator of a gene program controlling progenitor quiescence. Defining fundamental molecular mechanisms that render resident adipocyte progenitor cells quiescent holds promise to identify molecular barriers to adipogenesis and metabolically healthy fat.

脂肪组织扩张是由新脂肪细胞形成（导致增生性生长）和现有脂肪细胞肥大共同作用而发生的，其相对贡献可能会影响全身代谢健康。由于脂肪细胞发育失败而导致的先天性脂肪营养不良与严重的胰岛素抵抗有关。发育基因的遗传变异引起的脂肪生成障碍也容易导致胰岛素抵抗和 2 型糖尿病。这些例子反映了更多的数据，为代谢的假设提供了概念基础。健康取决于功能性脂肪细胞群。通过利用稳定同位素示踪剂和高精度质谱方法，我们现在还发现了脂肪生成的年龄依赖性下降，因此，脂肪细胞祖细胞（新脂肪细胞的细胞来源）必须存在于脂肪细胞中。尽管脂肪生成的下游驱动因素已得到很好的描述，但维持脂肪细胞祖细胞静止或其近端增殖转变的机制。为了解决这一知识空白，我们利用转录组数据以及新鲜分离的原代脂肪细胞祖细胞中可及染色质的基因组规模图来模拟负责静止状态的转录因子网络。表明孤儿核受体 NR4A1 是一个关键的调节节点体外和体内功能基因组学研究为我们以 NR4A1 为中心的假设提供了额外的支持：NR4A1 调节转录。这些数据还为两个相互关联的具体目标提供了理论基础，我们将测试 NR4A1 是脂肪细胞祖细胞静止的核心转录调节因子的假设。、CRISPRi 和 CRISPR/Cas9 在关键脂肪形成转录因子中功能性地询问 NR4A1-DNA 调控节点。我们将在基因组规模上对脂肪细胞祖细胞中 NR4A1 依赖性染色质调控景观进行公正的询问。我们将绘制 NR4A1-DNA 相互作用图谱，并将 NR4A1 与典型的顺式调控标记结合进行共定位。结合 NR4A1 功能的获得或丧失来绘制 NR4A1 依赖性染色质调控景观和转录输出的方法。总的来说，通过有针对性的功能基因组学实验和公正的基因组规模分析，我们将测试我们的 NR4A1 模型作为控制祖细胞静止的基因程序的主调节器，定义使驻留脂肪细胞祖细胞静止的基本分子机制有望识别脂肪生成的分子障碍。和代谢健康的脂肪。