Interaction of a 14-3-3 protein with the plant microtubule-associated protein EDE1

Abstract

The cell cycle-regulated protein ENDOSPERM DEFECTIVE 1 (EDE1) is a novel plant microtubule-associated protein essential for plant cell division and for microtubule organization in endosperm. EDE1 is only present on microtubules at mitosis and its expression is highly cell cycle regulated both at the protein and the transcript levels. To search for EDE1-interacting proteins, a yeast two-hybrid screen was used in which EDE1 was fused with GAL4 DNA binding domain and expressed in a yeast strain that was then mated with a strain carrying a cDNA library fused to the GAL4 transactivation domain. Candidate interacting proteins were identified and confirmed in vitro. 14-3-3 upsilon was isolated several times from the library screen. In in vitro tests, it also interacted with EDE1: 14-3-3 upsilon most strongly associates with EDE1 in its free form, but also weakly when EDE1 is bound to microtubules. This study shows that EDE1 is a cyclin-dependent kinase substrate but that phosphorylation is not required for interaction with 14-3-3 upsilon. The results suggest that 14-3-3 proteins may play a role in cytoskeletal organization of plant cells. The potential role of this interaction in the dynamics of EDE1 during the cell cycle is discussed.