HUMAN T LYMPHOCYTE FUNCTION IN LUPUS ERYTHEMATOSUS

我们描述了T细胞cAMP代谢紊乱， 系统性红斑狼疮（SLE）患者的淋巴细胞 我们最近发现的缺陷型cAMP依赖性 完整SLE T淋巴细胞中蛋白质的磷酸化表明 cAMP依赖性蛋白激酶（蛋白激酶A）疾病 本提案的目的是确定和描述 SLE T淋巴细胞蛋白激酶A功能异常 定量cAMP依赖性蛋白激酶活性， 酶的组成亚基，I型（RI）和 II（RII）调节和催化亚基（C亚基）。 具体 该建议的目的是（a）比较蛋白激酶A T的非颗粒和颗粒组分中的活性 淋巴细胞和T淋巴细胞亚群从活动和非活动 SLE受试者与对照。 cAMP依赖性蛋白激酶 活性将通过测量磷酸化 非颗粒和颗粒T细胞中组蛋白和Kemptide 萃取物(b)我们将部分纯化cAMP依赖性蛋白 SLE T淋巴细胞激酶的DEAE-Cellulose层析 为了确定异常RI、RII和/或C的存在， 亚单位。 异常的RI或RII亚基可以表现出cAMP的改变 结合，而异常C亚基可以具有降低的结合 对ATP的亲和力。 (c)蛋白激酶A亚单位的量 将通过ELISA和硝酸纤维素免疫标记进行定量， 确定是否存在一个或多个亚单位的缺陷， 解释蛋白激酶活性的改变 差异 激酶亚基的激酶活性和/或激酶量 将对T细胞亚群之间的差异进行定量，以确定是否存在免疫缺陷。 存在限制性激酶缺陷。 由于T淋巴细胞起着 在细胞免疫和体液免疫中起着不可或缺的作用， 重要的是要发现是否有缺陷的cAMP途径 部分解释了SLE中异常的免疫调节。 因此，我们的长期目标是确定缺陷蛋白质是否 激酶A功能导致抑制细胞活性异常。 蛋白激酶A缺陷的鉴定应提供 对这种自身免疫性疾病的发病机制的重要见解 disorder.

We have described a disorder of cAMP metabolism in T lymphocytes of subjects with systemic lupus erythematosus (SLE) Our recent identification of defective cAMP -dependent phosphorylation of proteins in intact SLE T lymphocytes suggests a disorder of cAMP -dependent protein kinase (Protein kinase A) The objective of this proposal is to identify and characterize the abnormal protein kinase A function in SLE T lymphocytes by quantifying cAMP -dependent protein kinase activities and the amounts of the enzyme's constituent subunits, the types I (RI) and II (RII) regulatory and catalytic subunits (C subunit). The specific aims of this proposal are (a) to compare protein kinase A activities in the nonparticulate and particulate fractions of T lymphocytes and T lymphocyte subsets from active and inactive SLE subjects with controls. cAMP -dependent protein kinase activity will be quantified by measuring the phosphorylation of histone and Kemptide in nonparticulate and particulate T cell extracts. (b) We will purify partially the cAMP -dependent protein kinases of SLE T lymphocytes by DEAE -cellulose chromatography in order to determine the presence of anomalous RI, RII and/or C subunits. Anomalous RI or RII subunits can exhibit altered cAMP binding while anomalous C subunits can possess reduced binding affinity for ATP. (c) The amounts of protein kinase A subunits will be quantified by ELISA and nitrocellulose immunolabeling to determine whether a deficiency of one or more subunits exists to explain altered protein kinase activities. Differences in the kinase activities and/or amounts of a kinase of a kinase subunit between T cell subsets will be quantified to determine whether a restricted kinase defect exists. Since the T lymphocyte plays an integral role in both cellular and humoral immunity, it is important to discover whether a defective cAMP pathway accounts, in part, for the aberrant immunoregulation in SLE. Thus, our longterm goal is to determine whether defective protein kinase A function results in abnormal suppressor cell activity. The identification of a protein kinase A defect should provide important insights into the pathogenesis of this autoimmune disorder.