STUDIES OF NATURALLY OCCURRING DEFECTS IN RESISTANCE TO INFECTION

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。所列机构为中心机构，不一定为研究者机构。原发性免疫缺陷疾病是研究最终导致有效免疫应答的分子事件的独特人类模型。在这笔赠款的支持下，我们的小组参与了与X连锁免疫缺陷疾病有关的基因的鉴定和这些基因产物的功能分析。对免疫紊乱的分子基础的理解极大地促进了受体-配体相互作用的概念，以及淋巴细胞活化、分化、信号转导和免疫球蛋白同种型转换的原理。将讨论三个具体目标。(1)Wiskott-Aldrich综合征（WAS）基因及其产物WAS蛋白的阐明。 我们是最近分离出WAS患者突变基因的团队的一员，WAS是一种大多数造血细胞谱系受到影响的疾病。我们收集的B和T细胞系来自50多个受影响的家庭成员，将允许一个代表性的突变分析的WAS，并将回答的问题，是否存在相关性之间的临床表型和突变类型。基于已知cDNA序列的氨基酸组成，我们概述了分析WAS蛋白质性质的策略。我们将专注于WAS基因的表达及其控制，其在细胞内的分布，WAS蛋白的物理和功能特性，将确定与WASP相互作用的其他蛋白质。(2)探讨T细胞转录激活缺陷在常见变异型免疫缺陷（CVI）患者中的作用。我们已经确定了一个亚组的CVI患者谁在共同的T细胞活化的广泛缺陷，这表明多种淋巴因子和膜结合的活化分子的转录可能是异常的。使用凝胶转移试验，我们将确定是否缺陷是由于异常结合的NF-AT或其他核因子相关的IL-2基因。如果证明结合减少，我们将寻找NF-AT复合物蛋白质的缺陷产生，并分析其在细胞质和核组分中的分布及其磷酸化状态。(3)探讨X连锁免疫缺陷患者基因治疗的可行性。我们已经开始与几位参与设计病毒载体的研究人员合作，这些病毒载体可以有效地将候选基因转移到靶细胞中，例如，T和B细胞系或来源于患者的干细胞。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Primary immunodeficiency diseases are unique human models to study the molecular events that ultimately lead to an efficient immune response. With the support of this grant, our group has participated in the identification of genes involved in X-linked immunodeficiency diseases and in the functional analysis of the products of these genes. The understanding of the molecular basis for immune disorders has contributed greatly to the concept of receptor-ligand interaction, and the principles of lymphocyte activation, differentiation, signal transduction and immunoglobulin isotype switching. Three specific aims will be addressed. (1) Elucidation of the gene responsible for the Wiskott-Aldrich syndrome (WAS) and its product, the WAS protein. We have been part of a team that recently isolated the gene mutated in patients with WAS, a disorder in which most hematopoietic cell lineages are affected. Our collection of B and T cell lines derived from members of over 50 affected families will permit a representative mutation analysis of WAS and will answer the question of whether a correlation exists between clinical phenotype and type of mutation. Based on the amino acid composition derived from the known cDNA sequence, we have outlined strategies to analyze the nature of the WAS protein. We will focus on WAS gene expression and its control, its distribution within the cell, the physical and functional properties of the WAS protein, will identify other proteins that interact with WASP. (2) Explore the role of defective transcriptional activation of T cells in patients with common variable immunodeficiency (CVI). We have identified a subgroup of CVI patients who have in common a broad defect of T cell activation, suggesting that the transcription of multiple lymphokines and membrane bound activation molecules may be abnormal. Using gelshift assays, we will determine if the defect is due to abnormal binding of NF-AT or other nuclear factors related to the IL-2 gene. If decreased binding is demonstrated, we will search for defective production of proteins of the NF-AT complex and analyze its distribution in the cytoplasm and in the nuclear fraction, and its phosphorylation state. (3) Explore the feasibility of gene therapy for patients with X-linked immunodeficiencies. We have initiated collaboration with several investigators involved in the design of viral vectors that efficiently transfer the candidate genes into target cells, e.g., T and B cell lines or stem cells derived from patients.