SIGNAL TRANSDUCTION MOLECULES ON MOUSE NK CELLS

自然杀伤（NK）细胞是一个大颗粒细胞亚群， 淋巴细胞（LGL）能够识别和裂解各种各样的 病毒感染或肿瘤靶细胞，以及某些 既往无致敏性或主要组织相容性的微生物 复合物（MHC）限制。 NK细胞不重排T细胞受体 基因，并且不表达细胞表面TcR分子或TcR基因的组分。 分化簇3（CD 3）转导复合物。 NK细胞确实 表达信号转导分子，包括CD 2、CD 16和CD 45。 然而，在这些中，只有CD 45不是NK细胞特异性的。 使用 针对大鼠IL-2激活的NK细胞的单克隆抗体， 最近鉴定了一种30 kD的蛋白，其在NK细胞上高度表达 细胞 这种蛋白质作为二硫键连接的二聚体发挥功能， 介导NK细胞中的跨膜信号传导。 此外，我们有 克隆并测序了编码这种新型信号转导的基因 老鼠体内的分子 由于其生物活性及其 在序列上的独特性，这种蛋白质可以被认为是一种新的 受体复合物，选择性地在NK细胞（NKR）上表达。 由于 该蛋白（PI）在NK细胞中可能具有的重要功能 介导的细胞毒活性，我们决定克隆小鼠NKR-P1 同源物 小鼠是一种研究更为广泛的动物模型， 让我们能够进行目前几乎 不可能在大鼠体内成功完成。 最近，由 将NKR-P1大鼠基因与存在于细胞中的小鼠cDNA交叉杂交， 从高度纯化的白细胞介素-2（IL-2）活化的， 小鼠自然杀伤（NK）细胞，我们成功地克隆了小鼠 作为cDNA和基因组基因两者的NKR-P1同源物。 三个类似 不同大小和顺序的信息在老鼠体内共同转录 NK细胞。 通过使用对每种共表达的 我们能够将这三个基因定位到一个共同的片段上， 与NK1.1被认为是 被映射。 编码NK1.1抗原的基因尚未被克隆。 对 在这些结果的基础上，我们现在认为我们处于 更准确地聚焦我们项目的目标。 这些目标将是：1） 为了研究小鼠NKR-P1基因家族的功能， 与NK1.1和NK2.1抗原的关系; 2）克隆启动子 来自基因组的所有三个共转录的小鼠NKR-P1基因的区域 克隆，我们已经分离，并试图表征一个transacting因子 所有这些都是共同的;和3）使用 不同大小的构建体，来源于可用的5'上游 区域片段，以更好地研究这些基因的调控。 我们 相信这些测试提供的信息将澄清 生理作用发挥y NKR-P1，并导致见解，以如何 利用其在体内的功能。

Natural killer (NK) cells are a subpopulation of large granular lymphocytes (LGL) that are able to recognize and lyse a wide variety of virally infected or neoplastic target cells, as well as certain microorganisms without previous sensitization or major histocompatibility complex (MHC)-restriction. NK cells do not rearrange T-cell receptor genes, and do not express cell surface TcR molecules or components of the cluster-of-differentiation 3 (CD3) transduction complex. NK cells do express signal transduction molecules, including CD2, CD16, and CD45. Among these, only CD45, however, is not NK cell specific. With the use of a monoclonal antibody against rat IL-2 activated NK cells, we have recently identified a 30 kD protein which is highly expressed on NK cells. This protein functions as a disulfide linked dimer capable of mediating transmembrane signalling in NK cells. Furthermore, we have cloned and sequenced the gene encoding this novel signal transduction molecule in the rat. Because of its biological activity and its uniqueness in sequence, this protein may be considered part of a new receptor complex,selectively expressed on NK cells (NKR). Due to the important function that this protein (PI) may have in the NK cell mediated cytotoxic activity, we decided to clone the mouse NKR-P1 homologue. The mouse is a more extensively studied animal model than the rat and allows us to perform experiments that are currently almost impossible to be successfully accomplished in the rat. More recently, by cross-hybridizing the NKR-P1 rat gene to the mouse CDNA present in a library generated from highly purified, interleukin-2 (IL-2) activated, mouse Natural Killer (NK) cells, we succeeded in cloning the mouse homologue of NKR-P1 as both CDNA and genomic genes. Three similar messages, differing in size and sequence, are co-transcribed in the mouse NK cells. By using probes specific for each of the co-expressed transcripts, we were able to map all 3 of these genes to a common segment of chromosome 6 closely linked to the region where NK1.1 is believed to be mapped. The gene encoding NK1.1 antigen has not yet been cloned. On the basis of these results, we now feel that we are in the position of focusing the aims of our project more precisely. These aims will be: 1) to study the function of mouse NKR-P1 gene family by first defining its relationship to the NK1.1 and NK2.1 antigens; 2) to clone the promotor region of all three co-transcribed mouse NKR-P1 genes from the genomic clones we have isolated and attempt to characterize a transacting factor common to all of them; and 3) to generate transgenic mice using constructs of different sizes, derived from the available 5' upstream region segments, to better study the regulation of these genes. We are confident that the information provided by these tests will clarify the physiological role played y NKR-P1 and lead to insights as to how to exploit its function in vivo.