Manservigi R, Gualandri R, Negrini M, Albonici L, Milanesi G, Cassai E, Barbanti-Brodano G
Institute of Microbiology, University of Ferrara, Italy.
Virology. 1988 Nov;167(1):284-8. doi: 10.1016/0042-6822(88)90080-3.
Expression of herpes simplex virus type 1 (HSV-1) glycoprotein B (gB-1) was obtained in human cells from the gB-1 gene cloned in the episomal replicating vector pBK-1, which contains the origin of replication and early region of the human papovavirus BK. Selective systems for the TK+ phenotype in TK-143B cells and for resistance to G418 in adenovirus 5-transformed 293 cells were used to obtain stable transformants that produced gB-1. While gB-1 expression in 143B cells required induction by HSV-1 early proteins, constitutive gB1 production was observed in 293 cells, where endogenous trans-acting factors probably replace the need for early viral products in the activation of the cloned gB-1 gene. The amount of recombinant gB-1 was comparable to that produced during HSV-1 lytic infection in human cells, due to amplification of the inserted gene in the replicating episomal vector. Expression of gB-1 was induced by cadmium and zinc when the promoter of the mouse metallothionein-I gene was placed upstream of gB1 structural sequences. The inducible system where the gB-1 gene is under the control of its own promoter could be employed to clarify the role of early viral products in induction of gB-1 synthesis. Constitutive expression of gB-1 in human cells could provide useful material for diagnostic purposes and for the preparation of a subunit vaccine against HSV infections.
在人细胞中,通过克隆于游离型复制载体pBK-1中的单纯疱疹病毒1型(HSV-1)糖蛋白B(gB-1)基因获得了gB-1的表达。该载体含有人类乳头瘤病毒BK的复制起点和早期区域。利用TK-143B细胞中TK+表型的选择系统以及腺病毒5转化的293细胞中对G418的抗性选择系统,获得了产生gB-1的稳定转化体。虽然143B细胞中gB-1的表达需要HSV-1早期蛋白诱导,但在293细胞中观察到了gB-1的组成型产生,内源性反式作用因子可能替代了早期病毒产物对克隆gB-1基因激活的需求。由于复制型游离载体中插入基因的扩增,重组gB-1的量与人细胞中HSV-1裂解感染期间产生的量相当。当小鼠金属硫蛋白-I基因的启动子置于gB-1结构序列上游时,gB-1的表达可被镉和锌诱导。可利用gB-1基因受其自身启动子控制的诱导系统来阐明早期病毒产物在gB-1合成诱导中的作用。gB-1在人细胞中的组成型表达可为诊断目的以及制备针对HSV感染的亚单位疫苗提供有用材料。