Topoisomerase I and II activities are required for Epstein--Barr virus replication

Abstract

The roles of topoisomerases I and II in Epstein-Barr virus (EBV) replication were investigated using Raji cells infected with EBV. The topoisomerase II inhibitor ellipticine inhibited the synthesis of EBV polypeptides at concentrations which did not affect total protein synthesis. Slot blot analysis of total cellular DNA showed that camptothecin and ellipticine inhibited replication of progeny EBV DNA in superinfected Raji cells at concentrations which did not inhibit synthesis of EBV early polypeptides prerequisite for EBV DNA replication. Analysis of the structure of EBV DNA termini demonstrated that both inhibitors affected the replicating EBV DNA. Gardella gel electrophoresis showed that both inhibitors affected the formation of the linear form of EBV DNA. However, restriction analysis of EBV DNA in superinfected Raji cells demonstrated that both inhibitors degraded neither endogenous nor exogenous EBV DNA. Cell viability was not affected by either inhibitor at the concentrations tested. These findings suggest that topoisomerase II is required for expression of the EBV genome and that both topoisomerases I and II are involved in replication of the EBV genome during the lytic phase of the life cycle. The effects of topoisomerase inhibitors on the circular form of EBV DNA during virus replication are discussed.Topoisomerases I and II are involved in gene expression and replication of eukaryotes (Maxwell & Gellert, 1985;Wang, 1985). In the case of animal viruses, topoisomerases play important roles in viral DNA replication, transcription, packaging and integration (Foglesong & Bauer, 1984;Snapka, 1986;Nishiyama et al., 1987;Benson & Huang, 1988;Champoux, 1988; Richter & Strausfeld, 1988;Ebert et al., 1990;Priel et al., 1990Priel et al., , 1991 Schaack et al., 1990a, b;Wong & Hsu, 1990;Yamada et al., 1990;Gu & Rhode, 1991;Priel et al., 1991 ;Wang & Rogler, 1991). Topoisomerase I changes the linking numbers of topological DNA domains in integral steps of one by introducing transient singlestrand breaks. Topoisomerase II breaks both DNA strands and resolves them, thereby changing topological linking numbers in steps of two.Recently, several specific inhibitors for mammalian DNA topoisomerases have been identified and characterized (Drlica & Franco, 1988). Camptothecin and ellipticine are specific inhibitors of mammalian topoisomerases I and II, respectively (Tewey et al., 1984;Hsiang et al., 1985). Camptothecin stablizes the covalent topoisomerase I-DNA complex, resulting in blockage of the rejoining step of the breakage-reunion reaction of topoisomerase I. Ellipticine affects the breakage-reunion reaction of topoisomerase II by stablizing the cleavable complex formed between topoisomerase II and DNA.These inhibitors are suitable for studying the functional involvement of topoisomerase activities in Epstein-Barr virus (EBV) gene expression and replication. EBV early antigen (EA) synthesis is induced in Raji cells by infection with P3HR-1 virus, followed by viral DNA replication and EBV late antigen synthesis. In ...

Kaposi's Sarcoma-Associated Herpesvirus ori - Lyt -Dependent DNA Replication: Involvement of Host Cellular Factors

“…But none of the herpesviruses encodes a DNArelaxing enzyme to release the topological stress created by DNA unwinding. Therefore, cellular DNA topoisomerases I and II are reportedly required for HSV-1, cytomegalovirus, and EBV lytic DNA replication (6,13,22). In this report, Topo I and II were found to bind to KSHV ori-Lyt DNA, suggesting their involvement in viral DNA replication.…”

Epstein-Barr Virus BGLF4 Kinase Induces Premature Chromosome Condensation through Activation of Condensin and Topoisomerase II

“…Specifically, Topo II␣ has been implicated as a critical enzyme involved in chromosome condensation and segregation (1,53). As demonstrated by inhibition experiments, Topo II␣ is required for HSV-1 and EBV replication (2,25), possibly in untangling concatemeric viral DNA. Here, we demonstrate that BGLF4 not only enhances Topo II-DNA interaction but also stimulates Topo II activity in a kinase activity-dependent manner ( Fig.…”

T cell proliferative response induced by DNA topoisomerase I in patients with systemic sclerosis and healthy donors.

“…In this regard, it has been shown recently that autoantibody to the p53 tumor suppressor protein was induced in mice immunized with complexes of murine p53 and SV40 large T antigen, but not in mice immunized with either protein separately (35). Topo I activity in host cells is known to be necessary for the replication and transcription of several viral genomes, such as adenovirus (36), SV40 (37,38), herpes simplex virus (39), EBV (40), and HIV (41). Since topo I binds tightly to viral proteins and/or DNA during viral replication and transcription (38,41), T cell autoimmunity may be initiated by the complex of topo I with certain viral proteins.…”