Importance of the Ser-132 phosphorylation site in cell transformation and apoptosis induced by the adenovirus type 5 E1A protein.
The 289-residue (289R) and 243R early region 1A (E1A) proteins of human adenovirus type 5 induce cell transformation in cooperation with either E1B or activated ras. Here we report that Ser-132 in both E1A products is a site of phosphorylation in vivo and is the only site phosphorylated in vitro by purified casein kinase II. Ser-132 is located in conserved region 2 near the primary binding site for the pRB tumor suppressor and, in 289R, just upstream of the conserved region 3 transactivation domain involved in regulation of early viral gene expression. Mutants containing alanine or glycine in place of Ser-132 interacted with pRB-related proteins at somewhat reduced efficiency; however, all Ser-132 mutants transformed primary rat cells in cooperation with E1B as well as or better than the wild type when both major E1A proteins were expressed. Such was not the case with mutants expressing only 289R. In cooperation with E1B, the Asp-132 and Gly-132 mutants yielded reduced numbers of smaller transformed foci. With activated ras, all Ser-132 mutants were significantly defective for transformation and the rare foci produced were small and contained extensive areas populated by low densities of flat cells. In the absence of E1B, all Ser-132 mutants induced p53-independent cell death more readily than virus expressing wild-type 289R. These results suggested that phosphorylation at Ser-132 may enhance the binding of pRB and related proteins and also reduce the toxicity of E1A 289R, thus increasing transforming activity.
Whalen
S G
SG
Department of Biochemistry, McGill University, Montreal, Quebec, Canada.
Marcellus
R C
RC
Barbeau
D
D
Branton
P E
PE
D000256:Adenoviridae / Q000235:genetics / Q000378:metabolism*; D017500:Adenovirus E1A Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D000595:Amino Acid Sequence; D017209:Apoptosis / Q000187:drug effects; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral / Q000187:drug effects; D015967:Gene Expression Regulation, Viral; D011905:Genes, ras; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism*
D017500:Adenovirus E1A Proteins; D012694:Serine