REGULATION OF CD45 ACTIVITY BY PHOSPHORYLATION

通过磷酸化调节 CD45 活性

基本信息

批准号：
6349854
负责人：
WALTER J ESSELMAN
金额：
$ 20.26万
依托单位：
MICHIGAN STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-02-01 至 2003-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6349854
关键词：
T lymphocyte biological signal transduction calcium flux enzyme activity gene mutation high performance liquid chromatography immunoprecipitation interleukin 2 mass spectrometry phosphorylation protein sequence protein tyrosine phosphatase radiotracer

项目摘要

DESCRIPTION (Adapted from Investigator's abstract): CD45 is a transmembrane receptor-type protein tyrosine phosphatase (PTP) that is highly expressed on virtually all cells of the hematopoietic lineage. Expression of CD45 is essential for antigen activation as well as for the differentiation of T and B lymphocytes. A key advantage to the study of CD45 is that the outcome of experiments modifying CD45 can be measured with a relevant functional assay - i.e., antigen activation. The investigator has recently reported the use of novel methods to identify the in vivo, naturally occurring phosphorylation sites of CD45. The hypothesis of this study is that the phosphorylation of CD45 regulates the function and the activity of CD45, and to relate CD45 phosphorylation to its function in antigen mediated activation. The investigator plans to address these goals as follows. The phosphorylation sites of CD45 in antigen activated cells will be identified by radiolabeling cells followed by isolation and analysis of CD45 by immunoprecipitation, peptide mapping, HPLC and MALDI-mass spectrometry. The role of key phosphorylation sties identified above in CD45 PTP activity will be determined by mutagenesis of phosphorylation sites followed by determination of enzyme activity and substrate specificity. Finally, the ability of phosphorylation site-mutant forms of CD45 to reconstitute the signaling responsiveness of CD45 negative cells will be determined. These studies are important to the understanding of phosphorylation events which regulate the receptivity of T cells to antigenic signals.

描述（根据调查员的摘要改编）：CD45是跨膜受体型蛋白酪氨酸磷酸酶（PTP）高度表达几乎所有造血谱系的细胞。 CD45的表达为对于抗原激活以及T和T的分化至关重要 B淋巴细胞。 CD45研究的关键优势是可以通过相关功能测定法测量修改CD45的实验 - 即抗原激活。调查员最近报告了鉴定体内，自然发生的新方法 CD45的磷酸化位点。这项研究的假设是 CD45的磷酸化调节CD45的功能和活性，将CD45磷酸化与抗原介导的功能相关联激活。调查人员计划解决这些目标，如下所示。这将鉴定出抗原活化细胞中CD45的磷酸化位点通过放射性标记的细胞，然后通过分离和分析CD45 免疫沉淀，肽映射，HPLC和MALDI-MAS光谱法。这关键磷酸化的作用是上述在CD45 PTP活性中的作用通过磷酸化位点的诱变，然后确定确定酶活性和底物特异性。最后， CD45磷酸化位点突显形式重建的能力将确定CD45负细胞的信号反应能力。这些研究对于理解磷酸化事件很重要调节T细胞对抗原信号的接受度。