CALCIUM AND POTASSIUM CHANNELS IN LYMPHOCYTE

本项目的主要目的是阐明T淋巴细胞中非基因组类固醇效应的机制。非基因组类固醇作用在过去十年中受到了广泛关注。它们的特点是起效迅速，不依赖于转录和蛋白质合成。非基因组类固醇作用是普遍存在的，并有据可查。我们和其他人已经证明类固醇激素会引起淋巴细胞中钙信号的快速变化。这些变化属于非基因组类固醇效应的范畴。在淋巴细胞中，TCR活化后的初始钙信号传导已显示通过两步过程发生，所述两步过程涉及IP 3介导的钙从内部储存释放和钙通过Ca 2+释放激活的Ca 2+通道流入。这种信号级联与脂筏中的酪氨酸激酶有关。脂筏，或洗涤剂不溶性微区，是表面膜成分，在淋巴细胞和许多其他细胞的信号转导和膜运输中起重要作用。我们建议，非基因组类固醇的行动介导的类固醇协会的蛋白质的脂筏。具体目标1：评价性腺类固醇对小鼠和人淋巴母细胞系细胞内游离Ca 2+水平的非基因组效应。特定目的2：确定淋巴母细胞系中是否存在小窝和/或洗涤剂不溶性微结构域（脂筏）。具体目标3：将淋巴细胞中的小窝和/或脂筏的存在与性腺类固醇对Ca 2+信号的非基因组作用相关联。本研究的结果将有助于我们了解信号转导、非基因组类固醇作用的细胞和亚细胞机制，以及免疫抑制机制。

The major goal of this project is to elucidate the mechanisms of non- genomic steroid effects in T lymphocytes. Non-genomic steroid actions have received much attention in the past decade. They are characterized by their rapid onset of action and by being independent of transcription and protein synthesis. Non-genomic steroid actions are ubiquitous and well documented. Steroid hormones have been shown by us and others to cause rapid changes in calcium signals in lymphocytes. These changes fall into the category of non- genomic steroid effects. In lymphocytes, initial calcium signaling upon TCR activation has been shown to occur by a two step process involving IP3 mediated release of calcium from internal stores and calcium influx through Ca2+ release-activated Ca2+ channels. This signaling cascade has been associated to tyrosine kinases in lipid rafts. Lipid rafts, or detergent insoluble microdomains, are surface membrane constituents that play important roles in signal transduction and membrane trafficking in lymphocytes and many other cells. We propose that non-genomic steroid actions are mediated by steroid association to the proteins of the lipid raft. The Specific Aims to be followed are: Specific Aim 1: Evaluate the non-genomic effects of the gonadal steroids on intracellular free Ca2+ levels in murine and human lymphoblastoid cell lines. Specific Aim 2: Determine the presence of caveolae and/or detergent insoluble microdomains (lipid rafts) in lymphoblastoid cell lines. Specific Aim 3: Correlate the presenc3e of caveolae and/or lipid rafts in l in lymphocytes with the non-genomic action of gonadal steroids on Ca2+ signaling. The results obtained in this project will contribute to our understanding of the cellular and subcellular mechanisms involved in signal transduction, non-genomic steroid action and for our understanding of the mechanisms involved in immunosuppression.