Phosphorylation-dependent regulation of nuclear import by 14-3-3

Abstract

14-3-3 proteins are phospho-serine/threonine binding proteins that play important roles in many regulatory processes including intracellular protein targeting. 14-3-3 proteins can anchor target proteins in the cytoplasm or the nucleus or mediate their nuclear export. So far no role for 14-3-3 in mediating nuclear import has been described. Myopodin is a dual compartment actin-bundling protein with tumor suppressor function in human bladder cancer. In muscle cells, myopodin redistributes between the nucleus and cytoplasm in a differentiation-dependent and stress-induced fashion. Here we show that importin a binding and subsequent nuclear import of myopodin is regulated by serine/threonine phosphorylation dependent binding of myopodin to 14-3-3. Furthermore, we establish that protein kinase A (PKA) and the calcium/calmodulin-dependent kinase II (CaMKII) phosphorylate the two 14-3-3 binding motifs of myopodin, thereby mediating the 14-3-3 binding and subsequent nuclear import of myopodin in myoblasts. In contrast, the protein phosphatase calcineurin dephosphorylates myopodin and abrogates the interaction between myopodin and 14-3-3. These results establish a novel paradigm for the promotion of nuclear import by 14-3-3 binding. They provide a molecular explanation for phosphorylation dependent nuclear import of nuclear localization signal containing cargo proteins.