Nuclear localization of protein kinase U-alpha is regulated by 14-3-3

14-3-3 proteins are intracellular, dimeric molecules that bind to and modif y the activity of several signaling proteins. We used human 14-3-3 zeta as a bait in the yeast two-hybrid system to screen a murine embryonic cDNA lib rary. One interacting clone was found to encode the carboxyl terminus of a putative protein kinase, The coding sequence of the human form (protein kin ase U alpha, PKU alpha) of this protein kinase was found in GenBank(TM) on the basis of sequence homology, The two-hybrid clone was also highly homolo gous to TOUSLED, an Arabidopsis thaliana protein kinase that is required fo r normal flower and leaf development. PKU alpha has been found by coimmunop recipitation to bind to 14-3-3 zeta in vivo. Our confocal laser immunofluor escence microscopic experiments revealed that PKU alpha colocalizes with th e cytoplasmic intermediate filament system of cultured fibroblasts in the G (1) phase of the cell cycle. PKU alpha is found in the perinuclear area of S phase cells and in the nucleus of late G(2) cells, Transfection of cells with a dominant negative form of 14-3-3 eta promotes the nuclear localizati on of PKU alpha. These results suggest that the subcellular localization of PKU alpha is regulated, at least in part, by its association with 14-3-3.