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

Abstract

14-3-3 proteins are intracellular, dimeric molecules that bind to and modify the activity of several signaling proteins. We used human 14-3-3ζ as a bait in the yeast two-hybrid system to screen a murine embryonic cDNA library. One interacting clone was found to encode the carboxyl terminus of a putative protein kinase. The coding sequence of the human form (protein kinase Uα, PKUα) of this protein kinase was found in GenBank(TM) on the basis of sequence homology. The two-hybrid clone was also highly homologous to TOUSLED, an Arabidopsis thaliana protein kinase that is required for normal flower and leaf development. PKUα has been found by coimmunoprecipitation to bind to 14-3-3ζ in vivo. Our confocal laser immunofluorescence microscopic experiments revealed that PKUα colocalizes with the cytoplasmic intermediate filament system of cultured fibroblasts in the G1 phase of the cell cycle. PKUα is found in the perinuclear area of S phase cells and in the nucleus of late G2 cells. Transfection of cells with a dominant negative form of 14-3-3η promotes the nuclear localization of PKUα. These results suggest that the subcellular localization of PKUα is regulated, at least in part, by its association with 14-3-3.