Abstract
Large T antigen (TAg) of the human polyomavirus JC virus (JCV) possesses DNA binding and helicase activities, which, together with various cellular proteins, are required for replication of the viral genome. We now show that JCV-infected cells expressing TAg accumulate in the G(2) phase of the cell cycle as a result of the activation of ATM- and ATR-mediated G(2) checkpoint pathways. Transient transfection of cells with a TAg expression vector also induced G(2) checkpoint signaling and G(2) arrest. Analysis of TAg mutants with different subnuclear localizations suggested that the association of TAg with cellular DNA contributes to the induction of G(2) arrest. Abrogation of G(2) arrest by inhibition of ATM and ATR, Chk1, and Wee1 suppressed JCV genome replication. In addition, abrogation of the G(2)-M transition by Cdc2 depletion disabled Wee1 depletion-induced suppression of JCV genome replication, suggesting that JCV replication is facilitated by G(2) arrest resulting from G(2) checkpoint signaling. Moreover, inhibition of ATM and ATR by caffeine suppressed JCV production. The observation that oligodendrocytes productively infected with JCV in vivo also undergo G(2) arrest suggests that G(2) checkpoint inhibitors such as caffeine are potential therapeutic agents for JCV infection.
Highlights
Large T antigen (TAg) of the human polyomavirus JC virus (JCV) possesses DNA binding and helicase activities, which, together with various cellular proteins, are required for replication of the viral genome
We found that G2 arrest in TAg-expressing cells was enhanced by the presence of a plasmid containing the JCV origin, indicating that viral DNA repcells, which are permissive for JCV replication, expressing JCV lication performed by TAg leads to further accumulation of early (TAg) or late (VP1) proteins 2 weeks after virus inoculation
We have shown that JCV TAg promotes viral DNA replication by arresting the cell cycle at the G2 phase as the result of induction of ataxia telangiectasia mutated (ATM)- and ATR-mediated G2 checkpoint pathways
Summary
JCV Large T Antigen Triggers G2 Arrest infected with SV40 or murine polyomavirus has been recently shown to activate the ataxia telangiectasia mutated (ATM)-mediated signaling pathway and thereby promote viral replication [11, 12]. It is not clear how polyomaviruses trigger DNA damage signaling. The Rad9-Rad1Hus (9:1:1) complex is recruited to RPA-coated ssDNA and brings the ATR activator, topoisomerase-binding protein-1 After their recruitment to sites of DNA damage, ATM and ATR phosphorylate various substrate proteins, including Chk and Chk, which in turn phosphorylate downstream factors to induce cell cycle arrest and facilitate DNA repair [16, 17]. We show that caffeine, an inhibitor of ATM and ATR function, markedly suppressed JCV replication and propagation after establishment of infection in host cells
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