Downregulation of cell-to-cell communication by the viralsrc gene is blocked by TMB-8 and recovery of communication is blocked by vanadate

Summary The viralsrc gene downregulates junctional communication, closing cell-to-cell membrane channels presumably by way of the phosphoinositide signal route. We show that TMB-8 [8-N, N-(diethylamino) octyl-3,4,5-trimethoxybenzoate] counteracts this downregulation in cells transformed by temperature-sensitive mutant Rous sarcoma virus: TMB-8 (36–72 m) raises junctional permeability when applied during activity ofsrc protein kinase, i.e., at steady permissive temperature; and TMB-8 inhibits the fall of junctional permeability, when the activity ofsrc protein kinase gets turned on. TMB-8 also (reversibly) inhibits the growth of the cells at permissive temperature and reverses the morphological changes associated with transformation. The morphological reversal lags several hours behind the junctional-permeability reversal. Communication recovers within a few minutes when the activity of thesrc protein kinase is turned off (in absence of TMB-8). Sodium orthovanadate (20 m) prevents this recovery, but it has no major effect on junctional permeability on its own. We discuss possible modes of action of these agents on critical stages of the signal route, related to intracellular Ca²⁺ and protein kinase C.     

利用人摄金蛋白(METALLOTHIONEIN)启动子和牛乳突瘤病毒在哺乳动物细胞中表达乙型肝炎表面抗原

用人Metallothioenin-Ⅱ启动子、乙型肝炎表面抗原基因,SV40早期基因的编接位点和多聚A位点构成了表达组件,然后插入到经改造过的BpV-1质粒中。所得质粒pdMTsAg-5转染小鼠C127细胞得到转化克隆。Ausria Ⅱ检测证明13株中有12株能产生HBsAg。对其中一株进行HBsAg收率观察,隔天收获为292.6～525.8μg/升,每天收获为200.9～369.0μg/升,可连续收获60天以上。经重金属离子诱导后,收率增至583～854.4μg/升。培养液经超滤浓缩和两次密梯离心后,可集中为一个狭窄的峰,顶峰的Ausria Ⅱ cpm为1.05×10~7,密度为1.20g/ml。