Function of a Putative Determinant in Hematopoiesis

造血作用中推定决定因素的功能

• 批准号: 7242617
• 负责人: JAMES J BIEKER
• 金额: $ 31.82万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-08-01 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7242617
• 关键词: Acetylation; Adult; Affect; Affinity Chromatography; Binding; Biological; Biological Process; Biology; Cell Nucleus; Cells; Chromatin; Chromatin Remodeling Factor; Complex; Cytoplasm; Data; Development; EP300 gene; Elements; Environment; Erythrocytes; Erythroid; Erythroid Cells; Erythropoiesis; Gene Expression Regulation; Genetic Transcription; Globin; HDAC1 gene; Hematopoiesis; Histone H3; In Vitro; Laboratories; Lighting; Modification; Molecular; Molecular Genetics; Nuclear; Play; Post-Translational Protein Processing; Process; Property; Proteins; Repression; Research Personnel; Role; SMARCA4 gene; Signal Transduction; Subcellular Fractions; System; Testing; Transcription Coactivator; Transcription Repressor/Corepressor; Transcriptional Activation; beta Globin; cell type; comparative; design; erythroid Kruppel-like factor; histone acetyltransferase; in vivo; mutant; novel; programs; promoter; transcription factor

DESCRIPTION (provided by applicant): Cell-restricted transcriptional modulators play a critical role in the process of selective gene regulation during hematopoiesis. We have been investigating the molecular and biological function of Erythroid Kruppel-Like Factor (EKLF). EKLF is a red cell-restricted transcription factor that is an essential component for completion of the erythroid program. Molecular and genetic data demonstrate that EKLF plays a major role in the developmental switch to adult beta-globin expression. It accomplishes this by means of its strong transcriptional activation function when bound to its cognate CACCC element at the beta-globin promoter, and by its ability to interact with coactivators that generate the proper chromatin configuration at the beta-like globin locus. Although the details of how EKLF integrates these signals remain to be fully deciphered, recent evidence unexpectedly suggests that it can function as a transcriptional repressor and may also play a role outside of the nucleus. Shifts between EKLF repression and activation are influenced by the cellular environment and by its interaction with corepressors. In addition, its modification by acetylation may also affect these decisions. This proposal is designed to expose the details of how these varied processes are controlled by identifying components of EKLF protein complexes and examining the role of post-translational modifications in these interactions. This will be accomplished by: (1) examining (a) in vitro and (b) italic how EKLF protein interactions with its activation partners are coordinated at the beta-globin promoter; (2) using tandem affinity purification to isolate EKLF protein complexes derived from selected cell types and subcellular fractions; (3) illuminating alternate roles for EKLF by (a) identifying EKLF repression targets and analyzing the role of its acetylation in repression and (b) characterizing cytoplasmic EKLF complexes.

描述（由申请人提供）：细胞限制的转录调节剂在造血过程中选择性基因调节过程中起关键作用。我们一直在研究红花样因子（EKLF）的分子和生物学功能。 EKLF是一种红细胞限制的转录因子，是完成红细胞程序的重要组成部分。分子和遗传数据表明，EKLF在发展到成人β-珠蛋白表达的发展中起主要作用。当与β-珠蛋白启动子处的同源CACCC元件绑定时，它通过其强大的转录激活函数来实现此目的，并通过与同时激活剂相互作用的能力，该共激活因子在类似Beta的Globin基因座上生成适当的染色质构型。尽管有关EKLF如何整合这些信号的细节仍待完全解密，但最近的证据出乎意料地表明，它可以作为转录阻遏物起作用，并且可能在核之外发挥作用。 EKLF抑制和激活之间的变化受细胞环境以及与核压子的相互作用的影响。此外，其通过乙酰化的修饰也可能影响这些决策。 该建议旨在通过识别EKLF蛋白复合物的组成部分来揭示如何控制这些多样的过程的细节，并检查后翻译后修饰在这些相互作用中的作用。这将通过：（1）检查（a）体外和（b）斜体蛋白与其激活伙伴的相互作用如何在β-珠蛋白启动子上协调； （2）使用串联亲和力纯化来分离源自选定的细胞类型和亚细胞级分的EKLF蛋白复合物； （3）通过（a）识别EKLF抑制靶标并分析其乙酰化在抑制作用中的作用和（b）表征细胞质EKLF复合物的作用。