Gene activation by varicella-zoster virus IE4 protein requires its dimerization and involves both the arginine-rich sequence, the central part, and the carboxyl-terminal cysteine-rich region

Varicella-zoster virus (VZV) open reading frame 4-encoded protein (IE4) pos sesses transactivating properties for VZV genes as well as for those of het erologous viruses. Since most transcription factors act as dimers, IE4 dime rization was studied using the mammalian two-hybrid system. Introduction of mutations in the IE4 open reading frame demonstrated that both the central region and the carboxyl-terminal cysteine-rich domain were important for e fficient dimerization. Within the carboxyl-terminal domain, substitution of amino acids encompassing residues 443-447 totally abolished dimerization. Gene activation by IE4 was studied by transient transfection with an IE4 ex pression plasmid and a reporter gene under the control of either the human immunodeficiency virus, type 1, long terminal repeat or the VZV thymidine k inase promoter. Regions of IE4 important for dimerization were also shown t o be crucial for transactivation. In addition, the arginine-rich domains Rb and Re of the amino-terminal region were also demonstrated to be important for transactivation, whereas the Ra domain as well as an acidic and bZIP-c ontaining regions were shown to be dispensable for gene transactivation. A nucleocytoplasmic shuttling of IE4 has also been characterized, involving a nuclear localization signal identified within the Rb domain and a nuclear export mechanism partially depending on Crm-1.