ROLE OF ETS1 IN LYMPHOID CELLS

ETS 1原癌基因在淋巴T细胞中表达水平较高， 细胞 ETS 1的表达与α-半乳糖苷酶的表达相关， T细胞受体（TCR α）在发育过程中。 ETS 1的表达是 维持在成熟的静息T细胞中，并且其水平在 activation. ETS 1是一种转录因子，与富含嘌呤的 基序（AGGAAGT）存在于许多不同基因的调控区， 也就是说，淋巴因子、TCR α、FOS、ETS 1、ETS 2。 为了理解 ETS 1基因产物在T细胞中，ETS 1活性被表达阻断， 反义ETS 1 RNA。 反义RNA能够与 正义RNA，从而阻止蛋白质的形成。 这些同基因的 然后将使用转染细胞系来研究 发生在ETS 1功能通路的下游。 在这方面，朱克特- 通过有限稀释将细胞亚克隆并用于转染。 的 全长ETS 1 cDNA和亚片段（代表ETS 1基因的5'区）， cDNA）克隆到基于巨细胞病毒的表达载体中，并在游离型中表达。 载体来表达反义RNA。 Jurkat细胞转染 这些表达载体通过电穿孔。 独立克隆人 G418中选择，并正在进一步调查中。 特别是， 和反映表面抗原表达模式的北方分析 （TCR α，TCR β，CD 3，CD 4），转染细胞的生长特性， 细胞（生长速度，肿瘤发生），这些细胞产生的淋巴因子 细胞（ELISA），以及通过免疫荧光法检测靶向的特定候选基因的调节。 ETS 1（使用北方分析和免疫沉淀和CAT测定）是 在一系列综合实验中进行。

The ETS1 proto-oncogene is expressed at a higher level in lymphoid T- cells. The expression of ETS1 is related to the expression of the alpha T-cell receptor (TCRalpha) during development. The expression of ETS1 is maintained in mature resting T cells, and its level decreases after activation. ETS1 is a transcription factor and binds to a purine-rich motif (AGGAAGT) found in the regulatory regions of many different genes, i.e., lymphokines, TCRalpha, FOS, ETS1, ETS2. To understand the function of ETS1 gene products in T-cells, the ETS1 activity is blocked by express- ing antisense ETS1 RNA. Antisense RNAs are able to form hybrids with sense RNA, thus blocking the formation of proteins. These isogenic transfectant cell lines will then be used to investigate the changes that occur downstream of ETS1 functional pathways. For this purpose, Jurkat T- cells were subcloned by limited dilution and used for transfection. The full-length ETS1 cDNA and subfragments (representing the 5' region of the cDNA) were cloned in cytomegalovirus-based expression vectors and episomal vectors to express antisense RNA. Jurkat cells were transfected with these expression vectors by electroporation. Independent clones were selected in G418 and are under further investigation. In particular, FACS and Northern analysis reflecting the expression pattern of surface antigen (TCRalpha, TCRbeta, CD3, CD4), the growth properties of the transfected cell (growth rate, tumorigenesis), the production of lymphokines by these cells (ELISA), and the modulation of specific candidate genes targeted by ETS1 (using Northern analysis and immunoprecipitation and CAT assays) are being performed in a series of comprehensive experiments.