A Lymphoblast Model for Diseases of Purine Metabolism

描述（由申请人提供）： 本实验室的长期目标是确定遗传原因，并阐明遗传性嘌呤代谢酶腺苷脱氨酶（ADA）缺乏症的生化和代谢影响。这些知识对于了解ADA缺乏症的临床表现的基础和开发有效的治疗方法至关重要，主要是严重联合免疫缺陷病（SCID），这是一种婴儿致命疾病（ADA缺乏症占所有SCID病例的15-20%）。然而，大约五分之一的ADA缺乏症患者更隐蔽地发展免疫缺陷，导致在儿童，青少年或成年后诊断。该实验室的主要目的之一是表征具有不同临床严重程度的患者中ADA基因突变的谱，并系统地定义特定突变对ADA的结构、功能和细胞表达的影响。这项研究对于确定患者的遗传组成（基因型）如何影响临床严重程度（表型）是必要的，并且它可以为预测预后以及可能对酶替代或基因治疗的反应提供基础。我们已经确定了一个重要的突变子集，干扰ADA折叠成一个积极的结构。我们现在将检查在淋巴细胞的细胞质中发现的伴侣蛋白的能力，以增强这些“折叠”突变的表达，这可能会调节免疫缺陷的严重程度。细胞外ADA可能调节腺苷对淋巴细胞功能的影响。我们已经确定了ADA与细胞膜蛋白CD 26的结合位点，并提供了证据表明ADA/CD 26相互作用对人体免疫功能不是必需的。我们现在将研究介导腺苷信号转导的ADA受体的直接相互作用，使用灵敏的生物化学方法检测ADA与从表达特定人腺苷受体亚型（但不包括CD 26或细胞内ADA）的细胞制备的膜的结合。我们还将表征CECR 1蛋白的腺苷和腺苷类似物的代谢中的催化活性和作用，CECR 1蛋白是一类昆虫分泌的“腺苷脱氨酶相关生长因子”的人类直系同源物，其被假定为控制腺苷的细胞外水平。

DESCRIPTION (provided by applicant): The long-term objectives of this laboratory are to define the genetic causes and elucidate the biochemical and metabolic effects of inherited deficiency of the purine metabolic enzyme adenosine deaminase (ADA). This knowledge is essential for understanding the basis of, and developing effective treatment for, the clinical manifestations of ADA deficiency, primarily Severe Combined Immunodeficiency Disease (SCID), a fatal disorder in infants (ADA deficiency accounts for 15-20% of all cases of SCID). However, about a fifth of patients with ADA deficiency develop immune deficiency more insidiously, leading to diagnosis later in childhood, adolescence, or adulthood. One of this laboratory's main aims has been to characterize the spectrum of ADA gene mutations in patients with different degrees of clinical severity, and to systematically define the effects of specific mutations on the structure, function, and cellular expression of ADA. This research is necessary to establish how a patient's genetic makeup (genotype) affects clinical severity (phenotype), and it may provide a basis for predicting prognosis and possibly the response to enzyme replacement or gene therapy. We have identified an important subset of mutations that interferes with the folding of ADA into an active structure. We will now examine the ability of chaperone proteins found in the cytoplasm of lymphoid cells to enhance the expression of these "folding" mutations, which may modulate the severity of immune deficiency. Extracellular ADA may regulate effects of adenosine on lymphocyte function. We have defined the binding site of ADA for the cell membrane protein CD26, and provided evidence that ADA/CD26 interaction is not essential for immune function in humans. We will now investigate the direct interaction of ADA receptors that mediate signal transduction by adenosine, using a sensitive biochemical method for detecting ADA binding to membranes prepared from cells that express specific human adenosine receptor subtypes (but not CD26 or intracellular ADA). We will also characterize the catalytic activity and role in the metabolism of adenosine and adenosine analogs of the CECR1 protein, which is the human ortholog of a class of insect secreted "adenosine deaminase related growth factors" postulated to control extracellular levels of adenosine.