Functional roles of lipids in early B cell receptor signaling.

描述（由申请人提供）：质膜脂质的物理性质在B细胞受体（BCR）激活中起着至关重要的作用，但关于脂质如何调节这一信号通路中蛋白质的组织和活性，我们知之甚少。克服这一基本知识差距对于阐明这一基本生物学功能的机制是必要的，并将导致免疫相关疾病的新干预措施的发展。本提案中概述的实验将确定质膜脂在调节BCR和免疫反应下游介质之间相互作用中的作用，并将开发调节细胞中脂质介导的相互作用的实验方法。工作假设是，健康的B细胞利用其膜脂的关键组成维持的异质性，平衡BCR，激活激酶和下调磷酸酶之间的相互作用，以促进早期激活步骤并抑制静息细胞中的受体活性。本研究通过定量测定正常B细胞和经定向脂质异质性扰动处理的细胞中BCR和质膜蛋白之间脂质介导的相互作用来验证这一假说。

DESCRIPTION (provided by applicant): The physical properties of plasma membrane lipids play vital roles in B cell receptor (BCR) activation, yet remarkably little is known regarding how lipids regulate the organization and activity of proteins within this signaling pathway. Overcoming this basic knowledge gap is necessary to elucidate the mechanisms underlying this essential biological function and will lead to the development of novel interventions for immune- related diseases. The experiments outlined in this proposal will identify the role plasma membrane lipids in regulating interactions between the BCR and down-stream mediators of the immune response, and will develop experimental methods to modulate lipid-mediated interactions in cells. The working hypothesis is that healthy B cells exploit heterogeneity maintained by a critical composition of its membrane lipids, to balance interactions between the BCR, activating kinases, and down-regulating phosphatases in order to facilitate early activation steps and suppress receptor activity in resting cells. The proposed research tests this working hypothesis by quantifying lipid-mediated interactions between the BCR and plasma membrane proteins in normal B cells and in cells treated with targeted perturbations of lipid heterogeneity. Guided by extensive preliminary data, two specific aims will be pursued: 1) Quantify and modulate effective interactions between the BCR and minimal membrane anchors and 2) Identify the roles of lipids in early B cell receptor activation. Both aims will utilize a new and tested method developed in the applicant's laboratory to directly measure interaction potentials between plasma membrane proteins using super-resolution imaging techniques. Both aims will also apply a novel class of membrane perturbations shown to modify the phase behavior of isolated plasma membrane vesicles and the composition of cross-linked BCR receptor clusters in cells. In the first aim, interaction energies will be measured between the BCR and membrane protein anchor motifs expressed in B cells and interactions will be modulated with biochemical perturbations. Under the second aim, the activity of full length proteins involved in BCR signaling will be measured and their effective interactions with other signaling components will be quantified over a range of experimental conditions. A predictive model will be developed that includes protein and lipid interactions, and plasma membrane heterogeneity will be investigated in lymphoma cells with known defects in BCR signaling. Although the proposed research will take place in B cells, it has implications for studying of role of lipids in other biological proceses and cell types. This approach is innovative because it draws on cutting edge experimental methodologies as well as the unique perspective that lipids impact functional processes by modulating effective interactions between embedded proteins. The proposed work is significant because it will establish a mechanism for lipid-mediated control of immune signaling processes enabling new strategies for the treatment of immune related disease through manipulation of plasma membrane physical properties.