Phosphorylation of polyoma T antigens
Phosphorylation of polyoma T antigens
复制标题
多瘤T抗原的磷酸化
DOI：
10.1016/0092-8674(79)90206-x
复制
发表时间：
1979
期刊：
影响因子：
64.5
通讯作者：
T. Benjamin
中科院分区：
文献类型：
--
作者：
B. Schaffhausen;T. Benjamin
The T antigens of polyoma virus have been examined for phosphoryiation in vivo and associated protein kinase activities in vitro. The 1DDK “large” T antigen is the major phosphoprotein among the T antigen species in vivo as determined by labeling virus-infected cells with 32P-orthophosphate. Hr-t mutants show normal phosphorylation of their 1 CIDK T antigens. The wild-type 50K plasma membraneassociated “middle” T antigen is also phosphorylated in the cell, but to a lesser extent than the 1 DDK; this low level phosphorylation is also observed in the presumably altered 56K protein induced by hrt mutant NG5g and in the 5DK truncated “middle” T of hr-t mutant SD15 Addition of dibutyryl cyclic AMP to the medium does not affect labeling of either large or middle T antigens in wild-type-or mutant-infected cells. Thus no differences are observed in T antigen phosphorylation in vivo between wild-type virus and hr-t mutants. Hr-t mutants are defective in a protein kinase activity assayed in vitro by adding y-32P-ATP to T antigen immunoprecipitates. In the case of wild-type virus, the 56K protein is the major phosphate acceptor in the in vitro kinase reaction, with a somewhat lower level of phosphoryiation observed in the 1DDK band. Hrt mutants NG5g and SD15 show no labeling of the altered 56K or 5DK, respectively, but do show detectable levels of 32P in the 1 DDK bands. A wild-type virus carrying a small deletion affecting the 1DDK and 56K bands shows a normal level of kinase activity associated with the truncated T antigens. Ts-a mutants appear to be normal with respect to the middle T antigen-associated kinase. Photoaffinity labeling of infected cell extracts with 6-arido cyclic AMP shows that the two major classes of regulatory subunits of cyclic AMP-dependent protein kinases are present in the immunoprecipltates. Phosphoryiatlon of histone Hi occurs when this substrate is added to immunoprecipitates of either mock-infected or virus-infected cells, again demonstrating the presence of cellular kinases. Further experiments will be required to determine whether the middle T antigen of polyoma virus is itself a protein kinase or simply a substrate for one or more cellular kinases.
