Vaccinia virus B1 kinase: phenotypic analysis of temperature-sensitive mutants and enzymatic characterization of recombinant proteins

Abstract

The vaccinia virus Bi gene encodes a 34-kDa protein with homology to protein kinases. In L cells infected nonpermissively with mutants containing lesions in the Bi gene (ts2 and ts25), the infectious cycle arrests prior to DNA replication. In this report, we demonstrate that DNA synthesis ceases when cultures infected with these mutants at 32°C are shifted to the nonpermissive temperature (39.5°C) in the midst of DNA replication. We also show that Bi protein is synthesized transiently during the early phase of infection, even when the progression to later stages of gene expression is prevented. Although wild-type (wt) Bl is stable, the ts Bi proteins are markedly labile in both L and BSC40 cells at both permissive and nonpermissive temperatures. These results suggest that the ts phenotype of the mutants is complex and may in part reflect a temperature-dependent requirement for kinase activity, an induction of temperature sensitivity in Bi substrates under nonpermissive

Inhibition of Vaccinia Virus Replication by Two Small Interfering RNAs Targeting B1R and G7L Genes and Their Synergistic Combination with Cidofovir

“…B1 is an essential serine/threonine protein kinase that is present in infecting virions (7,56) and is required for viral DNA synthesis, as shown by the phenotype of two temperature-sensitive mutants expressing a very labile B1 protein without kinase activity (11,55,56). The mechanism through which B1 contributes to DNA replication remains elusive, as the protein does not appear to phosphorylate any of the known components of the viral replication machinery (11).…”

SP600125 inhibits Orthopoxviruses replication in a JNK1/2 -independent manner: Implication as a potential antipoxviral

“…Taken together, questions still remain regarding the potential protein kinase(s) targeted by SP600125 during Orthopoxvirus infection causing the impairment of viral morphogenesis. Poxviruses encode two essential serine/threonine kinases, B1 (Traktman et al, 1989;Lin et al, 1992;Rempel and Traktman, 1992) and F10 (Lin and Broyles, 1994). While B1 plays a function during viral DNA replication (Traktman et al, 1989;Rempel et al, 1990;Domi and Beaud, 2000), F10 plays a role in the very early stages of virion morphogenesis (Wang and Shuman, 1995;Traktman et al, 1995).…”

Identification of sites phosphorylated by the vaccinia virus B1R kinase in viral protein H5R

“…Vaccinia virus is a large DNA virus that replicates in the host cell cytoplasm in granular sites called virosomes [ 1 ]. It encodes at least two protein kinases belonging to the cellular family of serine/threonine protein kinases, the products of the B1R [ 2 , 3 ] and F10L genes [ 4 ]. The F10L kinase is encapsidated in the virion and plays an essential role in virion morphogenesis [ 5 , 6 ].…”

“…The B1R protein kinase is expressed early in infection, is found in the virosomes, and is also packaged into virions [ 7 ]. It appears to be an essential viral protein, and temperature-sensitive mutations that map to the B1R gene produce virus that cannot replicate its DNA at the restrictive temperature [ 2 , 8 ]. The B1R kinase does not appear to have a broad substrate specificity, and, although it has some activity against the acidic protein, casein, this is a poor substrate compared with the enzyme's known physiological substrates.…”