Cyclin A1 directly interacts with B-myb and cyclin A1/cdk2 phosphorylate B-myb at functionally important serine and threonine residues: tissue-specific regulation of B-myb function

Cyclin A1 is tissue-specifically expressed during spermatogenesis, but it i s also highly expressed in acute myeloid leukemia (AML), Its pathogenetic r ole in AML and in the cell cycle of leukemic blasts is unknown. B-myb is es sential for G1/S transition and has been shown to be phosphorylated by the cyclin A2/cdk2 complex. Here it is demonstrated that cyclin A1 interacts wi th the C-terminal portion of B-myb as shown by glutathione S-transferase (G ST) precipitation. This interaction is confined to cyclin A1 because bindin g could not be detected between cyclin A2 and B-myb, Also, cdk2 was not pul led down by GST-8-myb from U937 lysates, In addition, co-immunoprecipitatio n of cyclin Al and B-myb in leukemic cells evidenced protein interaction in vivo. Baculovirus-expressed cyclin All cdk2 complexes were able to phospho rylate human as well as murine B-myb in vitro. Tryptic phosphopeptide mappi ng revealed that cyclin A1/cdk2 complexes phosphorylated the C-terminal par t of B-myb at several sites including threonine 447, 490, and 497 and serin e 581, These phosphorylation sites have been demonstrated to be important f or the enhancement of B-myb transcriptional activity. Further studies showe d that cyclin A1 cooperated with B-myb to transactivate myb binding site co ntaining promoters including the promoter of the human cyclin A1 gene. Take n together, the data suggest that cyclin A1 is a tissue-specific regulator of B-myb function and activates B-myb in leukemic blasts.