Mapping the organization of the peripheral nuclear compartment

绘制周围核区室的组织图

• 批准号: 9765209
• 负责人: Larry R. Gerace
• 金额: $ 58.09万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-01 至 2020-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9765209
• 关键词: 3-Dimensional; Address; Adipocytes; Biochemical; CRISPR/Cas technology; Cell Differentiation process; Cell Line; Cell Lineage; Cell Nucleus; Cells; Chemicals; Chromatin; Chromosome Structures; Chromosomes; Co-Immunoprecipitations; Crosslinker; Cultured Cells; Formulation; Future; Gene Expression; Genetic Transcription; Heterochromatin; Image; Light; Maps; Mass Spectrum Analysis; Mesenchymal Stem Cells; Methodology; Methods; Modeling; Muscle Cells; Nuclear; Nuclear Envelope; Nuclear Lamin; Nuclear Lamina; Nuclear Proteins; Nuclear Structure; Pathway interactions; Peripheral; Play; Polymers; Property; Protein Dynamics; Proteins; Proteomics; Regulation; Resolution; Role; Signal Transduction; Site; Specific qualifier value; Structure; Surface; Techniques; Work; Yeasts; adipocyte differentiation; cell type; chromatin protein; crosslink; embryonic stem cell; light microscopy; mechanical properties; protein protein interaction; public health relevance; stable isotope; stem cell differentiation; yeast two hybrid system

 DESCRIPTION (provided by applicant): Intranuclear compartments make essential contributions to chromosome structure and function. One such compartment is organized by the nuclear lamina (NL), a protein meshwork that lines the nucleoplasmic surface of the nuclear envelope (NE) in metazoan cells. The NL contains a polymeric assembly of nuclear lamins associated with a large number of transmembrane and peripheral proteins. In addition to serving as a structural framework for the NE, the NL modulates numerous signaling and transcriptional pathways, and provides a tethering and regulatory site for chromatin at the nuclear periphery. The nuclear lamina and associated peripheral heterochromatin can be viewed as comprising a distinctive nuclear compartment, herein termed the nuclear lamina/peripheral chromatin (NL/PC) compartment. Although various components of the NL have been described, the analysis has not been comprehensive. Moreover, approaches are underdeveloped for defining its compositional changes during cell differentiation, and for elucidating the protein-protein interactions that specify the NL network and its chromatin interactions. To address these fundamental issues, this project aims to develop methods to isolate sub-nuclear fractions enriched in the NL/PC compartment from mesenchymal stem cells (MSCs), and from myocytes and adipocytes differentiated from these cells. With the use of these fractions, systematic mass spectrometry (MS) methods will be developed to characterize the protein composition of the NL/PC compartment and to quantitatively chart its changes during differentiation. Chemical crosslinking (XL) combined with MS will be deployed to map the interactions of constituent proteins, and biochemical methods in tandem with super-resolution light microscopy will be used to validate these associations. From implementation of this complementary set of approaches, the project will provide a comprehensive reference map of the NL and the changes that it undergoes during differentiation of defined MSC lineages. This will potentiate the formulation of more general concepts on the functions of the NL/PC compartment, and will identify new components and nodes that can be targeted in future functional studies. This is expected to substantially advance our understanding of the role of the NL in cell-type-specific regulation of peripheral heterochromatin and gene expression.