A Proteome Map of Neutrophil Granules

中性粒细胞颗粒的蛋白质组图谱

基本信息

批准号：
6739106
负责人：
Kenneth R MCLEISH
金额：
$ 14.7万
依托单位：
UNIVERSITY OF LOUISVILLE
依托单位国家：
美国
项目类别：
财政年份：
2003
资助国家：
美国
起止时间：
2003-07-01 至 2005-10-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6739106
关键词：
biological signal transduction cell membrane clinical research collagenase cytotoxicity electrospray ionization mass spectrometry exocytosis granule high performance liquid chromatography human tissue kidney disorder matrix assisted laser desorption ionization membrane proteins neutrophil protein structure proteomics

项目摘要

DESCRIPTION (provided by applicant): Neutrophils contribute to health through their ability to kill invading microorganisms, however, these same mechanisms are used by neutrophils to produce injury in a number of acute renal diseases, including ischemic acute tubular necrosis, sepsis-induced acute renal failure, and acute glomerulonephritis. Participation in these diseases requires a phenotypic change from benign circulating neutrophils to cells capable of extensive release of toxic oxygen radicals and proteolytic enzymes, a process termed priming. We showed recently that neutrophil priming is dependent on exocytosis of intracellular granules. Thus, defining the mechanisms of exocytosis will likely lead to new approaches to prevention and treatment of several devastating renal diseases. Neutrophils contain four different types of intracellular granules, each of which is characterized by a particular set of luminal and membrane proteins. Understanding the molecular mechanisms of exocytosis is hindered by the lack of knowledge of the proteins associated with each granule. This problem is amenable to the application of proteomic techniques. This application proposes to develop proteome maps of the intracellular granule and plasma membranes. Membranes from azurophil, specific, and gelatinase granules and plasma membranes will be isolated from human neutrophils. A major challenge to using a proteomic approach to define membrane proteins is their poor solubility due to hydrophobicity. The solubility problem impairs extraction, separation, and identification of these proteins. We propose to extract membrane-associated and integral membrane proteins through application of sequential solubilization techniques. The proteins will then be identified using two-dimensional electrophoresis and mass spectrometry, combined with informatics. This knowledge will allow us to formulate hypotheses related to the molecular mechanisms and signal transduction pathways that control neutrophil exocytosis and priming. These hypotheses will lead to NIH applications to confirm these mechanisms and to identify methods to interrupt neutrophil participation in acute renal injury.

描述（由申请人提供）：中性粒细胞通过杀死入侵微生物的能力对健康做出贡献，然而，中性粒细胞也利用这些相同的机制对许多急性肾病造成损伤，包括缺血性急性肾小管坏死、脓毒症引起的急性肾病。衰竭和急性肾小球肾炎。参与这些疾病需要从良性循环中性粒细胞到能够大量释放有毒氧自由基和蛋白水解酶的细胞的表型变化，这一过程称为启动。我们最近表明，中性粒细胞启动依赖于细胞内颗粒的胞吐作用。因此，明确胞吐作用的机制可能会带来预防和治疗几种破坏性肾脏疾病的新方法。中性粒细胞含有四种不同类型的细胞内颗粒，每种颗粒都具有一组特定的管腔蛋白和膜蛋白。由于缺乏对与每个颗粒相关的蛋白质的了解，理解胞吐作用的分子机制受到阻碍。这个问题可以通过应用蛋白质组学技术来解决。该申请旨在开发细胞内颗粒和质膜的蛋白质组图谱。将从人嗜中性粒细胞中分离出天青粒细胞膜、特异性细胞膜和明胶酶颗粒膜以及质膜。使用蛋白质组学方法来定义膜蛋白的一个主要挑战是它们由于疏水性而溶解度差。溶解度问题损害了这些蛋白质的提取、分离和鉴定。我们建议通过应用顺序溶解技术来提取膜相关蛋白和整合膜蛋白。然后，将使用二维电泳和质谱并结合信息学来鉴定蛋白质。这些知识将使我们能够制定与控制中性粒细胞胞吐作用和启动的分子机制和信号转导途径相关的假设。这些假设将导致 NIH 申请确认这些机制并确定中断中性粒细胞参与急性肾损伤的方法。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Comparison of proteins expressed on secretory vesicle membranes and plasma membranes of human neutrophils.

人中性粒细胞分泌囊泡膜和质膜上表达的蛋白质的比较。

DOI：
发表时间：
2008-04-15
期刊：
影响因子：
0
作者：
Uriarte, Silvia M;Powell, David W;Luerman, Gregory C;Merchant, Michael L;Cummins, Timothy D;Jog, Neelakshi R;Ward, Richard A;McLeish, Kenneth R
通讯作者：
McLeish, Kenneth R

Counterregulation of clathrin-mediated endocytosis by the actin and microtubular cytoskeleton in human neutrophils.

人中性粒细胞中肌动蛋白和微管细胞骨架对网格蛋白介导的内吞作用的反调节。

DOI：
发表时间：
2009-04
期刊：
影响因子：
0
作者：
Uriarte, Silvia M;Jog, Neelakshi R;Luerman, Gregory C;Bhimani, Samrath;Ward, Richard A;McLeish, Kenneth R
通讯作者：
McLeish, Kenneth R

A proteomic screen identified stress-induced chaperone proteins as targets of Akt phosphorylation in mesangial cells.

蛋白质组学筛选将应激诱导的伴侣蛋白鉴定为系膜细胞中 Akt 磷酸化的靶标。

DOI：
发表时间：
2006-07
期刊：
影响因子：
4.4
作者：
Barati, Michelle T;Rane, Madhavi J;Klein, Jon B;McLeish, Kenneth R
通讯作者：
McLeish, Kenneth R

骨髓相关蛋白 14 是人中性粒细胞中的 p38 MAPK 底物。

DOI：
发表时间：
2005-06-01
期刊：
影响因子：
0
作者：
Lominadze, George;Rane, Madhavi J;Merchant, Michael;Cai, Jian;Ward, Richard A;McLeish, Kenneth R
通讯作者：
McLeish, Kenneth R

Heat shock protein 27 regulates neutrophil chemotaxis and exocytosis through two independent mechanisms.

热休克蛋白 27 通过两种独立的机制调节中性粒细胞趋化性和胞吐作用。

DOI：
发表时间：
2007-02-15
期刊：
影响因子：
0
作者：
Jog, Neelakshi R;Jala, Venkatakrishna R;Ward, Richard A;Rane, Madhavi J;Haribabu, Bodduluri;McLeish, Kenneth R
通讯作者：
McLeish, Kenneth R