Kolesnitchenko V, Snart R S
Département de Physiologie Cellulaire, Université Pierre et Marie Curie, Paris, France.
AIDS Res Hum Retroviruses. 1992 Dec;8(12):1977-80. doi: 10.1089/aid.1992.8.1977.
Within the long terminal repeat (LTR) of the human immunodeficiency virus type 1 (HIV-1) provirus there exists a steroid hormone-responsive element corresponding to the TGTTCT sequence identified as the glucocorticoid receptor binding element within the LTR of mouse mammary tumor virus. We have used an LTR(HIV-1)-chloramphenicol acetyl transferase (CAT) plasmid construct to transfect infected H9V3 and noninfected H9 cells. Four hours before harvest the cells were divided into two parts and half was treated with hydrocortisone (10(-7) M). The cells were harvested and washed, and the CAT activity was measured. In eight repeat experiments an increased expression of the CAT gene has consistently been observed in H9V3 cells in response to the glucocorticoid but no significant effect of the steroid was observed in noninfected cells. Double transfection of LTR(HIV-1)-TAT and LTR(HIV-1)-CAT into noninfected H9 cells results in a cell population in which the CAT gene was responsive to glucocorticoid stimulation. A time course and dose response for the steroid effect have been determined and the binding of steroid receptor fo the LTR-DNA characterized by gel retardation experiments.
在人类免疫缺陷病毒1型（HIV-1）前病毒的长末端重复序列（LTR）中，存在一个类固醇激素反应元件，其对应于被鉴定为小鼠乳腺肿瘤病毒LTR内糖皮质激素受体结合元件的TGTTCT序列。我们使用了一种LTR（HIV-1）-氯霉素乙酰转移酶（CAT）质粒构建体来转染受感染的H9V3细胞和未感染的H9细胞。收获前4小时，将细胞分成两部分，一半用氢化可的松（10^(-7) M）处理。收获并洗涤细胞，然后测量CAT活性。在八次重复实验中，始终观察到H9V3细胞中CAT基因的表达因糖皮质激素而增加，但在未感染的细胞中未观察到类固醇的显著作用。将LTR（HIV-1）-TAT和LTR（HIV-1）-CAT双重转染到未感染的H9细胞中，产生了一个细胞群体，其中CAT基因对糖皮质激素刺激有反应。已经确定了类固醇作用的时间进程和剂量反应，并通过凝胶阻滞实验对LTR-DNA的类固醇受体结合进行了表征。
