Functional proteomics in differentiating erythroid cells

分化红细胞的功能蛋白质组学

基本信息

批准号：
8460913
负责人：
JORG BUNGERT
金额：
$ 24.83万
依托单位：
UNIVERSITY OF FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-05-15 至 2016-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8460913
关键词：
Adult Affect Anemia Binding Biochemical Biological Assay Biotin Biotinylation Cells Chromatin Chromatin Loop Chromosomes Code Complex DNA DNA Sequence Data Development Disease Elements Enhancers Erythroid Erythroid Cells Fetal Liver Future Gene Expression Gene Expression Regulation Gene Targeting Genes Genetic Transcription Globin Goals Hemoglobinopathies Human Knowledge Life Locus Control Region MEL Gene Mass Spectrum Analysis Mediating Methodology Modeling Molecular Molecular Conformation Mus Mutation Population Precipitation Proteins Proteomics Recruitment Activity Regulation Regulatory Element Role Sampling Sickle Cell Anemia Site Staging Streptavidin Technology Thalassemia Transcription factor genes Transcriptional Activation Transgenic Mice USF2 gene Undifferentiated Yolk Sac chromatin immunoprecipitation chromatin protein cofactor crosslink embryonic stem cell gain of function human GATA1 protein insight loss of function magnetic beads novel promoter protein complex protein crosslink protein function public health relevance research study restriction enzyme transcription factor

项目摘要

DESCRIPTION (provided by applicant): The human ?-globin gene locus consists of five genes that are expressed in a developmental stage- and erythroid-specific manner and regulated by a locus control region (LCR) located far upstream of the genes. Mutations in the ?-globin gene locus are quite frequent in the human population and associated with mild to severe anemias. Most of these mutations are located within the coding region of the adult ?-globin gene or within regulatory sequences affecting expression of the adult gene. Among the most prominent of ?-globin associated diseases are sickle cell anemia (SCA) and ?- thalassemias. Current treatments for these diseases are unsatisfactory and much effort is being invested in developing novel forms of therapies. We anticipate that further knowledge of how the globin genes are activated during development will create new opportunities for the treatment of hemoglobinopathies. For example, recent evidence suggests that the regulatory elements of the globin locus, including the LCR and the gene promoters, come in close proximity during activation of gene expression. If it is understood how genes are brought into close proximity to the LCR it may be possible in the future to modulate these interactions in such a way that associations of the ?-globin genes are favored over that of the adult ?-globin gene in definitive erythroid cells. We propose to perform a comprehensive analysis of the composition and function of protein complexes containing transcription factors involved in globin gene regulation. In these studies we will utilize novel methodology, including expression of biotin tagged proteins in differentiating erythroid cells and in transgenic mice, to examine the role of transcription factors and associated co-factors in the regulation of chromatin accessibility, globin locus conformational changes, and recruitment of transcription complexes. Biotinylated transcription factors will be isolated from cells using streptavidin coated magnetic beads and subjected to mass-spectrometry for identifying associated proteins, to chromatin immunoprecipitation (ChIP) for identifying associated DNA sequences, and to a combined ChIP and chromosome conformation capture (3C) assay for identifying proteins that function in the context of a globin locus configuration in which the LCR and promoters are in close proximity.

描述（由申请人提供）：人类？ - 斑蛋白基因基因座由五个基因组成，这些基因以发育阶段和红细胞特异性方式表达，并由位于基因上游较远的基因座控制区域（LCR）调节。 - 珠蛋白基因基因座的突变在人群中很常见，并且与轻度至重度贫血有关。这些突变中的大多数位于成年？ - 珠蛋白基因的编码区域内或影响成年基因表达的调节序列。在？ - 珠蛋白相关疾病中最突出的是镰状细胞贫血（SCA）和 - thalassemias。这些疾病的当前治疗方法不令人满意，并且在开发新型疗法上投入了很多努力。我们预计，在开发过程中如何激活球蛋白基因将为血红蛋白病治疗创造新的机会。例如，最近的证据表明，在基因表达激活过程中，球蛋白基因座的调节元件（包括LCR和基因启动子）在接近附近。如果了解如何使基因与LCR密切接近，那么将来有可能以这种相互作用的方式调节这些相互作用，以至于 - 格珠蛋白基因的关联比在确定性红细胞中的成年基因相对于成年基因的相关性。我们建议对蛋白质复合物的组成和功能进行全面分析，这些蛋白质复合物含有球蛋白基因调节涉及的转录因子。在这些研究中，我们将利用新方法，包括在分化红细胞和转基因小鼠中表达生物素标记的蛋白质，以检查转录因子和相关的co剂在调节染色质可及性中的作用，Globin lobus stous构象变化，转录复合物的骨骼骨骼。 Biotinylated transcription factors will be isolated from cells using streptavidin coated magnetic beads and subjected to mass-spectrometry for identifying associated proteins, to chromatin immunoprecipitation (ChIP) for identifying associated DNA sequences, and to a combined ChIP and chromosome conformation capture (3C) assay for identifying proteins that function in the context of a globin locus configuration in which the LCR and promoters are紧邻。

项目成果

期刊论文数量（5）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Distinct and overlapping DNMT1 interactions with multiple transcription factors in erythroid cells: Evidence for co-repressor functions.

DOI：
10.1016/j.bbagrm.2016.09.007
发表时间：
2016-12
期刊：
Biochimica et biophysica acta
影响因子：
0
作者：
Dimitris N. Papageorgiou;Elena Karkoulia;Alexandra Amaral-Psarris;P. Burda;Katarzyna E. Kolodziej;J. Demmers;J. Bungert;T. Stopka;J. Strouboulis
通讯作者：
Dimitris N. Papageorgiou;Elena Karkoulia;Alexandra Amaral-Psarris;P. Burda;Katarzyna E. Kolodziej;J. Demmers;J. Bungert;T. Stopka;J. Strouboulis

GATA-1 genome-wide occupancy associates with distinct epigenetic profiles in mouse fetal liver erythropoiesis.

DOI：
10.1093/nar/gkt167
发表时间：
2013-05
期刊：
Nucleic acids research
影响因子：
14.9
作者：
Papadopoulos GL;Karkoulia E;Tsamardinos I;Porcher C;Ragoussis J;Bungert J;Strouboulis J
通讯作者：
Strouboulis J

GATA1 and PU.1 Bind to Ribosomal Protein Genes in Erythroid Cells: Implications for Ribosomopathies.