STUDIES OF REGULATION IN E.COLI EXTENDED TO LYMPHOMAS

我们项目的目的仍然是了解糖皮质激素的作用。 激素对基因表达的影响。我们正在调查两种反应： 小鼠T淋巴瘤细胞的裂解及纤维连接蛋白的诱导 纤维肉瘤细胞系。 到目前为止，我们对糖皮质激素诱导的淋巴细胞溶解的研究已经完成。 与小鼠淋巴样细胞系。近几个月来，我们延长了我们的 对人急性淋巴细胞白血病细胞系CEM的作用。我们有 在其亚克隆中检测了糖皮质激素耐药的基础 CEM-C1，已知含有功能性糖皮质激素受体。 DNA脱甲基剂5-aza-2‘对CEM-C1细胞的作用 脱氧胞苷高频产生糖皮质激素敏感亚克隆 (约10%)。这表明DNA甲基化是 用于失活所需的“裂解”功能的表达 这些细胞对糖皮质激素敏感。 我们已经跟进了我们的观察结果，糖皮质激素增加了10- HT1080纤维肉瘤细胞纤维连接蛋白生物合成速率提高20倍。 用一个人纤维连接蛋白的cDNA克隆来探测Northern blots，我们发现 该探针与高分子量的信使核糖核酸 地塞米松处理的HT1080。这种信使核糖核酸的水平要低得多。 在未经处理的HT1080细胞中，在CEM细胞中检测不到，CEM细胞不 产生纤维粘连蛋白。通过对细胞总RNA的“槽”印迹分析，我们 发现激素后纤维连接蛋白RNA水平增加了16到32倍 治疗。此外，这种诱导被糖皮质激素抑制。 对抗者，RU486。本研究表明，地塞米松诱导的 纤维连接蛋白生物合成的刺激是由于 糖皮质激素受体介导的纤维连接蛋白基因积聚。 目前正在进行研究，以检查这种影响是否归因于 转录速度的提高。(D)

The aim of our project remains to understand effects of glucocorticoid hormones on gene expression. We are investigating two responses: the lysis of T-lymphoma cells and the induction of fibronectin in a fibrosarcoma cell line. Our studies of glucocorticoid-induced lymphocytolysis have been done so far with murine lymphoid cell lines. In recent months we have extended our work to the human acute lymphoblastic leukemic cell line CEM. We have examined the basis for glucocorticoid resistance in a subclone of that line, CEM-C1, known to contain functional glucocorticoid receptor. Treatment of CEM-C1 cells with the DNA-demethylating agent 5-aza-2' deoxycytidine yielded glucocorticoid-sensitive subclones at high frequency (approximately 10%). This indicates that DNA methylation is responsible for inactivating the expression of a "lytic" function required for glucocorticoid-sensitivity in these cells. We have followed up on our observation that glucocorticoids increase 10-\to 20-fold the rate of fibronectin biosynthesis in HT1080 fibrosarcoma cells. Using a human fibronectin cDNA clone to probe Northern blots, we found that this probe hybridizes with a high molecular weight mRNA from dexamethasone-treated HT1080. This mRNA is present at a much lower level in untreated HT1080 cells and is undetectable in CEM cells, which do not produce fibronectin. By "slot" blot analysis of total cellular RNA, we found a 16-\to 32-fold increase in fibronectin RNA level after hormone treatment. Moreover, this induction is inhibited by a glucocorticoid antagonist, RU486. This study demonstrates that the dexamethasone-induced stimulation of fibronectin biosynthesis is the result of an increase in accumulated fibronectin mRNA mediated by the glucocorticoid receptor. Studies are currently in progress to examine whether this effect is due to an increased rate of transcription. (D)