Agnoprotein-1a of avian polyomavirus budgerigar fledgling disease virus: identification of phosphorylation sites and functional importance in the virus life-cycle

The avian polyomavirus budgerigar fledgling disease virus (BFDV) encodes an unusual set of four agnoproteins in its late upstream region. Of the two p airs of these proteins, which overlap each other in two different reading f rames, the p(L1)-promoted agnoprotein-1a (agno-1a) is the dominant species and is able to support virus propagation in the absence of the other three polypeptides. Viral BFDV agno-1a, and also agno-1a expressed via an influen za virus vector, consists of a complex series of electrophoretically separa ble subspecies that can be reduced by phosphatase action down to a primary unphosphorylated protein with an apparent molecular mass of 31 kDa. Through peptide mass spectrometry and site-directed mutagenesis, the positions of four serine and three threonine residues have been determined as phosphate- accepting groups, which are partially modified by the combined action of th ree different cellular kinases. Since extensively phosphorylated agno-1a is required for its intracellular function, control over VP protein expressio n, and unphosphorylated agno-1a is observed as an additional component in t he BFDV virion, both extreme subspecies appear to be drawn from that comple x mixture, which also includes the intermediate stages of phosphorylation.