Phosphorylation of large tumour antigen by cdc2 stimulates SV40 DNA replication

Abstract

Simian virus 40 large tumour antigen (T) is a replication origin binding protein required for viral DNA synthesis. Unphosphorylated T antigen is deficient in promoting DNA replication in vitro but can be activated by phosphorylation at residue threonine 124 by the cdc2 protein kinase. This observation demonstrates that T is regulated by phosphorylation and provides a model for cdc2 function in the control of DNA replication.

Nuclear protein phosphorylation and growth control

“…Phosphorylation at this residue may depend on prior phosphorylation at Ser-120 to generate a recognition determinant (Flotow et al, 1990;Scheidtmann et al, 199 la;Umpress et al, 1992), but the kinase which phosphorylates Ser-120 is not known. Thr-124 is phosphorylated by the cell cycle-regulated kinase cdkl (p34cdc2) in vitro (McVey et al, 1989) and Ser-677 has recently been shown to be phosphorylated in vitro by dsDNA-PK (Lees-Miller et al, 1990;Chen et al, 1991). Protein kinase activities which target other phosphorylation sites in T antigen have yet to be identified.…”

“…Further evidence for control of replication by phosphorylation came from two different approaches. In the first, T antigen expressed in bacterial cells was shown to be inactive in replicating SV40 DNA in vitro (McVey et al, 1989). However, when the bacterially expressed protein was phosphorylated at Thr-124 (by cdkl in vitro), it acquired a hyperactive role in replication (i.e.…”

“…However, when the bacterially expressed protein was phosphorylated at Thr-124 (by cdkl in vitro), it acquired a hyperactive role in replication (i.e. more active than T antigen from HeLa cells, which is phosphorylated at this and at other sites) and bound extremely efficiently to site II of the replication origin (McVey et al, 1989). These findings indicated a positive role for phosphorylation of T antigen at Thr-124 and suggested that phosphorylation at another site(s) downregulates activity.…”

SV40 T antigen interacts with Nbs1 to disrupt DNA replication control

“…Replication of SV40 DNA requires a single viral protein, SV40 large T antigen (T), and the cellular DNA replication machinery (Challberg and Kelly 1989;Stillman 1989;Hurwitz et al 1990). Although the replication initiation function of T is partially regulated by cell-cycle-dependent phosphorylation (McVey et al 1989;Prives 1990), SV40-infected cells escape the strict cellular control of one round of DNA replication per cell cycle and massively amplify the viral genome (Tegtmeyer 1972;Chou et al 1974;Botchan et al 1979). Meanwhile, SV40 induces more than one round of cellular DNA replication within a given cell cycle and regularly induces endoreduplication and hyperploidy.…”

Casein kinase II phosphorylates bovine papillomavirus type 1 E1 in vitro at a conserved motif.

“…The biochemical activities and DNA replication functions of E1 resemble those of the simian virus 40 large T antigen (Clertant & Seif, 1984 ;Fanning, 1992 ;Fanning & Knippers, 1992 ;Seif, 1984). T antigen is known to be phosphorylated at multiple sites in vivo in a manner that regulates the DNA replication and transformation activities of the protein (Chen & Paucha, 1990 ;Fanning & Knippers, 1992 ;McVey et al, 1989 ;Prives, 1990 ;Schneider & Fanning, 1988). The locations and functions of phosphorylation sites on E1 are less well characterized than those of T antigen.…”